Metabolic dysregulation of planktonic bacteria

Abstract

Compositions comprising 2-hydroxycarboxylic acid for dysregulating the metabolism of planktonic bacteria and their use. This disclosure provides compositions for dysregulating the metabolism of planktonic bacteria, the compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid. This disclosure further provides compositions for dysregulating the metabolism of planktonic bacteria, the compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid and antimicrobial compounds. Dysregulation of the metabolism of planktonic bacteria can be measured, among other examples, by a decrease in the MIC of concomitant antimicrobial agents, an enhancement of in vivo antimicrobial effects, or aerobic respiration of metabolically active cells (reduction of non-fluorescent resazurin to highly fluorescent resorphine), which can lead to sensitization of planktonic bacteria to antimicrobial compounds and / or disruption of resistance of planktonic bacteria to antimicrobial compounds.

Description

[Technical Field]

[0001] This disclosure relates to compositions comprising preferred enantiomers of 2-hydroxycarboxylic acids for dysregulation of the metabolism of planktonic bacteria and sensitization of planktonic bacteria to antimicrobial compounds, and to methods for dysregulating the metabolism of planktonic bacteria and sensitizing planktonic bacteria to antimicrobial compounds using the above-described compositions. [Background technology]

[0002] In nature, bacteria can alternate between two main modes of reproduction: a unicellular life phase in which cells are free-swimming (planktonic) and a multicellular life phase in which cells are sessile and grow within colonies such as biofilms. In the unicellular life phase, bacteria can disperse and form colonies in new environments, while biofilms allow for a more persistent and cooperative mode of survival that is advantageous for the survival and sustained growth of sessile cells. In this alternating cycle, bacteria achieve two physiological transitions through differential gene expression: (i) transition from planktonic bacteria to sessile cells within a biofilm, and (ii) transition from sessile cells to new planktonic bacteria.

[0003] Attached bacteria differ metabolically from planktonic bacteria due to physiological characteristics expressed to aid survival within physicochemical conditions and constrained environments (such as biofilms). While estimates vary, some suggest that up to approximately 40% of bacterial genes may be upregulated or downregulated in the transition from planktonic to biofilm and vice versa. Furthermore, bacteria can develop and deploy resistance mechanisms to treatments such as antibiotics and biocides, potentially impairing therapeutic and decontamination efforts. Resistance mechanisms deployed by attached biofilm bacteria may differ from those deployed by planktonic bacteria.

[0004] The presence of planktonic bacteria can be harmful in healthcare, drinking water distribution systems, food, and marine industries. For example, in the food industry, pathogenic bacteria can be present inside processing facilities, causing food spoilage and endangering consumer health. In hospital settings, resistant and non-resistant planktonic bacteria have also been shown to persist on medical equipment surfaces, patient tissues, or as airborne or fluid-derived single-cell bacteria causing persistent infections. Furthermore, planktonic bacteria contribute to life-threatening infections and diseases in humans or animals, such as sepsis, waterborne or foodborne diarrheal diseases, urinary tract infections, lung infections, otitis media, periodontitis, sinusitis, chronic wounds, eye infections, sexually transmitted infections, bacterial meningitis and osteomyelitis, and vaginitis. If these bacteria are resistant to disinfectants, serious health management and contamination problems arise.

[0005] There is a need to eliminate difficult-to-remove bacterial infections that exist throughout the natural environment, including human diseases, crop maintenance, commercial animal farms, animal diseases, and biological contamination in marine organisms, membranes, sensors and piping, wind turbines, and other applications.

[0006] Antimicrobial resistance can reduce the effectiveness of the therapeutic agents applied, which may be antibiotics or agents with different modes of antimicrobial action, such as bacteriophages, disinfectants, saline solutions, chlorine-based compounds (such as bleach), iodine-based compounds, copper-based compounds, and heavy metals, and are used to help eliminate airborne bacteria.

[0007] Bacterial biofilms are complex communities of bacteria that adhere to surfaces and / or to each other, held together by a self-producing polymer matrix composed primarily of polysaccharides, secreted proteins, and extracellular DNA. Biofilm formation can begin with the attachment of free-floating planktonic microorganisms to a surface. Once colonization begins, the biofilm grows through a combination of cell division and recruitment. Biofilm formation and other forms of bacterial persistence can be avoided if planktonic bacteria can be removed or killed before a biofilm develops. Furthermore, since biofilms can release individual bacterial cells that can settle in other spaces where new biofilms may form, it would be advantageous to suppress, destroy, or kill released bacteria before they can form new biofilms.

[0008] There is a need for alternative treatments to aid in the destruction, suppression, and / or elimination of planktonic bacteria, or at least to provide sensitizers to complement additional bacterial treatments, which may prevent or disrupt the development of resistance mechanisms developed by the bacteria. This disclosure seeks to provide improved or alternative methods for the destruction, suppression, and elimination of planktonic bacteria, in particular methods that can be used in combination with antimicrobial compounds such as antibiotics or biocides.

[0009] The foregoing description of the background technology is intended solely to facilitate understanding of this disclosure. The discussion is not an acknowledgment or recognition that any of the materials referred to were, or were part of, common general knowledge as of the priority date of the application. [Overview of the project] [Means for solving the problem]

[0010] Summary of the Invention This disclosure provides a composition for dysregulating the metabolism of planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid.

[0011] This disclosure further provides compositions for dysregulating the metabolism of planktonic bacteria, the compositions comprising preferred enantiomers of 2-hydroxycarboxylic acids and antimicrobial compounds.

[0012] Metabolic dysregulation of planktonic bacteria can be measured, among other things, by a decrease in the MIC of concomitant antimicrobial agents, an enhancement of in vivo antimicrobial effect, or aerobic respiration of metabolically active cells (reduction of non-fluorescent resazurin to highly fluorescent resorphine), which can lead to sensitization of planktonic bacteria to antimicrobial compounds and / or the breakdown of resistance to antimicrobial compounds. When an antimicrobial compound is introduced to planktonic bacteria either while a preferred enantiomer of 2-hydroxycarboxylic acid is dysregulating the metabolism of planktonic bacteria (preferably resulting in sensitization) or after a preferred enantiomer of 2-hydroxycarboxylic acid has dysregulated the metabolism of planktonic bacteria (preferably resulting in sensitization), the metabolic dysregulation of planktonic bacteria can lead to enhanced bacterial killing or enhanced mitigation of infection. Metabolic dysregulation of planktonic bacteria can be measured as metabolic suppression, metabolic enhancement, or metabolic suppression followed by metabolic enhancement.

[0013] The Disclosure further provides compositions for dysregulating the metabolism of planktonic bacteria that are resistant to one or more antimicrobial compounds (i.e., planktonic bacteria that are susceptible to these antimicrobial compounds and referred to as “non-resistant planktonic bacteria”), the compositions comprising a preferred enantiomer of 2-hydroxycarboxylic acid, the preferred enantiomer of 2-hydroxycarboxylic acid, which induces dysregulation of the metabolism of planktonic bacteria and sensitizes the bacteria to antimicrobial compounds.

[0014] The present disclosure further provides a composition for dysregulating the metabolism of planktonic bacteria (referred to as "resistant planktonic bacteria") that are resistant to one or more antimicrobial compounds, the composition comprising a preferred enantiomer of 2-hydroxycarboxylic acid, the preferred enantiomer of 2-hydroxycarboxylic acid inducing dysregulation of the metabolism of planktonic bacteria and sensitizing the bacteria to antimicrobial compounds.

[0015] The present disclosure further provides a composition for sensitizing planktonic bacteria to antimicrobial compounds. The composition comprises a preferred enantiomer of 2-hydroxycarboxylic acid, and the preferred enantiomer of 2-hydroxycarboxylic acid induces dysregulation of the metabolism of planktonic bacteria.

[0016] The present disclosure further provides a composition for sensitizing planktonic bacteria to antimicrobial compounds, the composition comprising a preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, and the preferred enantiomer of 2-hydroxycarboxylic acid induces dysregulation of the metabolism of planktonic bacteria.

[0017] The present disclosure further provides a composition for enhancing the efficacy of an antimicrobial compound for decontaminating a surface comprising planktonic bacteria, the composition comprising a preferred enantiomer of 2-hydroxycarboxylic acid, and the preferred enantiomer of 2-hydroxycarboxylic acid induces dysregulation of the metabolism of planktonic bacteria.

[0018] The present disclosure further provides a composition for enhancing the efficacy of an antimicrobial compound for decontaminating a surface comprising planktonic bacteria, the composition comprising a preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, and the preferred enantiomer of 2-hydroxycarboxylic acid induces dysregulation of the metabolism of planktonic bacteria.

[0019] The antimicrobial compound in the composition of the present invention may be added together with the preferred enantiomer of 2-hydroxycarboxylic acid, or may be added after the preferred enantiomer of 2-hydroxycarboxylic acid has been introduced into the planktonic bacteria. If necessary, when the antimicrobial compound is added after the preferred enantiomer of 2-hydroxycarboxylic acid, the planktonic bacteria are sensitized to the antimicrobial compound by the preferred enantiomer of 2-hydroxycarboxylic acid.

[0020] In the composition of the present invention, if necessary, dysregulation of the metabolism of planktonic bacteria results in sensitization of the bacteria to the antimicrobial compound. In the composition of the present invention, if necessary, dysregulation of the metabolism of planktonic bacteria is suppression of the metabolism of planktonic bacteria, or suppression of bacterial metabolism followed by enhancement of bacterial metabolism.

[0021] The planktonic bacteria can be resistant planktonic bacteria, non-resistant planktonic bacteria, or a combination of resistant planktonic bacteria and non-resistant planktonic bacteria.

[0022] The composition of the present disclosure may contain 0.001% to 100% of the total 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient, and / or other active agents. In one aspect, the proportion (%) of the less preferred enantiomer of 2-hydroxycarboxylic acid in the composition is less than 20%.

[0023] The antimicrobial compound can be a bactericidal, bacteriostatic, antibiofilm or disinfectant antimicrobial compound. The antimicrobial compound can be an antibiotic, or an agent having a different mode of antimicrobial action, such as a bacteriophage, a disinfectant, physiological saline, a chlorine-based compound (such as a bleach), an iodine-based compound, a copper-based compound, a heavy metal, etc.

[0024] Compositions of the present disclosure comprising preferred enantiomers of 2-hydroxycarboxylic acid may be used to dysregulate the metabolism of free-flowing planktonic bacteria on biological and non-biological surfaces, and in fluids and gases (including air), and, if necessary, to sensitize the planktonic bacteria to antimicrobial compounds.

[0025] This disclosure further provides a method for dysregulating the metabolism of planktonic bacteria, and this method is i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to floating bacteria.

[0026] This disclosure further provides a method for dysregulating the metabolism of planktonic bacteria, and this method is i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, a preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0027] This disclosure further provides a method for sensitizing planktonic bacteria to an antimicrobial compound, the method being i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0028] This disclosure further provides a method for sensitizing planktonic bacteria to an antimicrobial compound, the method being i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0029] This disclosure further provides a method for enhancing the efficacy of antimicrobial compounds for decontaminating surfaces containing airborne bacteria, the method being described as follows: i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0030] This disclosure further provides a method for enhancing the efficacy of antimicrobial compounds for decontaminating surfaces containing airborne bacteria, the method being described as follows: i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0031] This invention provides a method for treating or preventing bacterial infections caused by airborne bacteria, and this method is i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to the infection site, Here, preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0032] This disclosure provides a method for treating or preventing bacterial infections caused by airborne bacteria, and this method is i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to the infection site, Here, a preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound, and the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria.

[0033] The Disclosure further provides compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid for use in treating or preventing bacterial infections, wherein the bacterial infections are caused by planktonic bacteria, and the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacteria.

[0034] The Disclosure further provides compositions comprising a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound for use in treating or preventing an infection in a subject, wherein the infection is caused by a planktonic bacterium, and the preferred enantiomer of the 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacterium.

[0035] The Disclosure further provides compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid for use in treating or preventing bacterial infections in a subject, wherein the bacterial infection is caused by planktonic bacteria, and the compositions sensitize the planktonic bacteria to an antimicrobial compound, wherein the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacteria.

[0036] The Disclosure further provides compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid for use in combination with an antimicrobial compound to treat or prevent an infection in a subject, wherein the infection is caused by a planktonic bacterium, the composition sensitizes the planktonic bacterium to the antimicrobial compound, and the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacterium.

[0037] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the preparation of compositions for dysregulating the metabolism of planktonic bacteria.

[0038] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the preparation of compositions for dysregulating the metabolism of planktonic bacteria in combination with antimicrobial compounds.

[0039] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for metabolic dysregulation in planktonic bacteria.

[0040] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds for metabolic dysregulation of planktonic bacteria.

[0041] The disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acid to sensitize planktonic bacteria to antimicrobial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0042] The disclosure further provides the use of a preferred enantiomer of 2-hydroxycarboxylic acid to sensitize planktonic bacteria to an antimicrobial compound, wherein the preferred enantiomer of 2-hydroxycarboxylic acid is combined with the antimicrobial compound, and the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0043] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acid in the preparation of compositions for sensitizing planktonic bacteria to antimicrobial compounds, wherein preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0044] This disclosure provides the use of a preferred enantiomer of 2-hydroxycarboxylic acid in the preparation of a composition for sensitizing planktonic bacteria to an antimicrobial compound, wherein the preferred enantiomer of 2-hydroxycarboxylic acid is combined with the antimicrobial compound, and the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria.

[0045] In the above use, if necessary, dysregulation of the metabolism of planktonic bacteria is suppression of the metabolism of planktonic bacteria.

[0046] The preferred enantiomer of the 2-hydroxycarboxylic acid may have the same composition as the antimicrobial compound, or a different composition. The antimicrobial compound may be a bactericidal, bacteriostatic, antibiofilm, or disinfectant antimicrobial compound. The antimicrobial compound may be a bactericidal or bacteriostatic antibiotic, or an agent having a different mode of antimicrobial action, such as a bacteriophage, antibiofilm, quorum sensing inhibitor, disinfectant, saline solution, chlorine-based compounds (such as bleaches), iodine-based compounds, copper-based compounds, or heavy metals.

[0047] This disclosure provides a kit for inhibiting the metabolism of planktonic bacteria, and this kit is, a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes the instruction manual.

[0048] This disclosure provides a kit for inhibiting the metabolism of planktonic bacteria, and this kit is, a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes instructions for use, Here, a preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0049] This disclosure provides a kit for sensitizing planktonic bacteria to an antimicrobial compound, and this kit is a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes instructions for use, Here, preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0050] This disclosure provides a kit for sensitizing planktonic bacteria to an antimicrobial compound, and this kit is a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes instructions for use, Here, the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0051] In the methods, uses, and kits of the present invention, the antimicrobial compound may be added together with the preferred enantiomer of 2-hydroxycarboxylic acid, or added after the preferred enantiomer of 2-hydroxycarboxylic acid has been introduced to the planktonic bacteria. If the antimicrobial compound is added after the preferred enantiomer of 2-hydroxycarboxylic acid as needed, resistant and / or non-resistant planktonic bacteria are sensitized to the antimicrobial compound by the preferred enantiomer of 2-hydroxycarboxylic acid.

[0052] In the methods, uses, and kits of the present invention, dysregulation of the metabolism of planktonic bacteria, if necessary, leads to bacterial sensitization to antimicrobial compounds. In the methods, uses, and kits of the present invention, dysregulation of the metabolism of planktonic bacteria, if necessary, is suppression of the metabolism of planktonic bacteria, or suppression of bacterial metabolism followed by enhancement of bacterial metabolism.

[0053] Planktonic bacteria may be resistant planktonic bacteria, non-resistant planktonic bacteria, or a combination of resistant and non-resistant planktonic bacteria.

[0054] In the embodiments described above, a preferred enantiomer of 2-hydroxycarboxylic acid is D-lactic acid or a pharmaceutically acceptable salt thereof. [Brief explanation of the drawing]

[0055] Further features of this disclosure are described more fully in the following description of some non-limiting embodiments thereof. This description is included for illustrative purposes only and should not be understood as a limitation to the broad overview, disclosure, or description of the above disclosure. The following description will be made with reference to the accompanying drawings. [Figure 1A-B]Figures 1A and 1B are graphs showing the results of adding D-lactate at 2-fold serial dilutions to suspension cultures of mixed-phenotypic P. aeruginosa clinical isolates. Figures 1A, 1C, and 1E show measurements of culture absorbance as an indicator of suspension bacterial density. As can be seen from each histogram, the treatment does not result in a significant decrease in culture absorbance across the evaluated D-lactate concentration range. However, Figures 1B, 1D, and 1F show that when metabolic rate (reduction of resazurin to fluorescent resorphine) is used as a readout, a breakpoint occurs at D-lactate concentrations between 0.25 mg / mL and 0.5 mg / mL within the same treatment range. Figure 1A shows the absorbance (culture density) of P. aeruginosa clinical isolates exhibiting smooth colony morphology (n=8) after treatment at the indicated D-lactate concentrations. Figure 1B shows the relative fluorescence measured after adding resazurin to the same P. aeruginosa clinical isolates (n=8) with smooth colony morphology. The breakpoint in the metabolism of these cultures is evident between treatment with D-lactate at 0.25 mg / mL to 0.5 mg / mL. Figure 1C shows the absorbance (culture density) of P. aeruginosa clinical isolates with coarse colony morphology (n=5) after treatment with the indicated D-lactate concentration range. Figure 1D shows the relative fluorescence measured after adding resazurin to the same P. aeruginosa clinical isolates (n=5) with coarse colony morphology. The breakpoint in the metabolic response of these cultures is evident between treatment with D-lactate at 0.25 mg / mL to 0.5 mg / mL. Figure 1E shows the absorbance (culture density) of P. aeruginosa clinical isolates (n=11) with mucoid colony morphology after treatment with the indicated D-lactate concentration range. Figure 1F shows the relative fluorescence measured after adding resazurin to clinical isolates of P. aeruginosa (n=11) with the same mucoid colony morphology. The breakpoint in the metabolic response is evident during D-lactate treatment between 0.25 mg / mL and 0.5 mg / mL. [Figure 1C-D]Figures 1A and 1B are graphs showing the results of adding D-lactate at 2-fold serial dilutions to suspension cultures of mixed-phenotypic P. aeruginosa clinical isolates. Figures 1A, 1C, and 1E show measurements of culture absorbance as an indicator of suspension bacterial density. As can be seen from each histogram, the treatment does not result in a significant decrease in culture absorbance across the evaluated D-lactate concentration range. However, Figures 1B, 1D, and 1F show that when metabolic rate (reduction of resazurin to fluorescent resorphine) is used as a readout, a breakpoint occurs at D-lactate concentrations between 0.25 mg / mL and 0.5 mg / mL within the same treatment range. Figure 1A shows the absorbance (culture density) of P. aeruginosa clinical isolates exhibiting smooth colony morphology (n=8) after treatment at the indicated D-lactate concentrations. Figure 1B shows the relative fluorescence measured after adding resazurin to the same P. aeruginosa clinical isolates (n=8) with smooth colony morphology. The breakpoint in the metabolism of these cultures is evident between treatment with D-lactate at 0.25 mg / mL to 0.5 mg / mL. Figure 1C shows the absorbance (culture density) of P. aeruginosa clinical isolates with coarse colony morphology (n=5) after treatment with the indicated D-lactate concentration range. Figure 1D shows the relative fluorescence measured after adding resazurin to the same P. aeruginosa clinical isolates (n=5) with coarse colony morphology. The breakpoint in the metabolic response of these cultures is evident between treatment with D-lactate at 0.25 mg / mL to 0.5 mg / mL. Figure 1E shows the absorbance (culture density) of P. aeruginosa clinical isolates (n=11) with mucoid colony morphology after treatment with the indicated D-lactate concentration range. Figure 1F shows the relative fluorescence measured after adding resazurin to clinical isolates of P. aeruginosa (n=11) with the same mucoid colony morphology. The breakpoint in the metabolic response is evident during D-lactate treatment between 0.25 mg / mL and 0.5 mg / mL. [Figure 1E-F]Figures 1A and 1B are graphs showing the results of adding D-lactate at 2-fold serial dilutions to suspension cultures of mixed-phenotypic P. aeruginosa clinical isolates. Figures 1A, 1C, and 1E show measurements of culture absorbance as an indicator of suspension bacterial density. As can be seen from each histogram, the treatment does not result in a significant decrease in culture absorbance across the evaluated D-lactate concentration range. However, Figures 1B, 1D, and 1F show that when metabolic rate (reduction of resazurin to fluorescent resorphine) is used as a readout, a breakpoint occurs at D-lactate concentrations between 0.25 mg / mL and 0.5 mg / mL within the same treatment range. Figure 1A shows the absorbance (culture density) of P. aeruginosa clinical isolates exhibiting smooth colony morphology (n=8) after treatment at the indicated D-lactate concentrations. Figure 1B shows the relative fluorescence measured after adding resazurin to the same P. aeruginosa clinical isolates (n=8) with smooth colony morphology. The breakpoint in the metabolism of these cultures is evident between treatment with D-lactate at 0.25 mg / mL to 0.5 mg / mL. Figure 1C shows the absorbance (culture density) of P. aeruginosa clinical isolates with coarse colony morphology (n=5) after treatment with the indicated D-lactate concentration range. Figure 1D shows the relative fluorescence measured after adding resazurin to the same P. aeruginosa clinical isolates (n=5) with coarse colony morphology. The breakpoint in the metabolic response of these cultures is evident between treatment with D-lactate at 0.25 mg / mL to 0.5 mg / mL. Figure 1E shows the absorbance (culture density) of P. aeruginosa clinical isolates (n=11) with mucoid colony morphology after treatment with the indicated D-lactate concentration range. Figure 1F shows the relative fluorescence measured after adding resazurin to clinical isolates of P. aeruginosa (n=11) with the same mucoid colony morphology. The breakpoint in the metabolic response is evident during D-lactate treatment between 0.25 mg / mL and 0.5 mg / mL. [Figure 2]Figure 2 is a graph showing that D-lactate enhances the suppression of metabolic activity in the presence of ceftriaxone. A: S. aureus 29213 (MSSA). B: S. aureus 510 (MRSA). Relative metabolic activity is shown in black for ceftriaxone monotherapy and in gray for ceftriaxone + A: 10 mM D-lactate or B: 0.1 mM D-lactate. [Figure 3] Figure 3 is a graph showing that D-lactate enhances the suppression of metabolic activity in the presence of meropenem. A: S. aureus 29213 (MSSA). B: S. aureus 510 (MRSA). Relative metabolic activity is shown in black for meropenem monotherapy and in gray for meropenem + 10 mM D-lactate. [Figure 4] Figure 4 is a graph showing that D-lactate enhances the suppression of metabolic activity in the presence of methicillin. A: S. aureus 29213 (MSSA). B: S. aureus 510 (MRSA). Relative metabolic activity is shown in black for methicillin monotherapy and in gray for methicillin + 10 mM D-lactate. [Figure 5] Figure 5 is a graph showing that D-lactate enhances the suppression of metabolic activity in the presence of rifampicin. A: S. aureus MSSA293 (MSSA). B: S. aureus 510 (MRSA). Relative metabolic activity is shown in black for rifampicin monotherapy and in gray for rifampicin + 0.16 mM D-lactate. [Figure 6] Figure 6 is a graph showing that D-lactate enhances the suppression of metabolic activity in S. aureus 510 (MRSA) in the presence of tetracycline. Relative metabolic activity is shown in black for rifampicin monotherapy and in gray for tetracycline + 10 mM D-lactate. [Figure 7]Figure 7 is a graph showing that D-lactate enhances the suppression of metabolic activity in the presence of vancomycin. A: S. aureus 29213 (MSSA). B: S. aureus 510 (MRSA). Relative metabolic activity is shown in black for vancomycin monotherapy and in gray for vancomycin + A: 10 mM D-lactate or B: 1.0 mM D-lactate. [Figure 8] Figure 8 is a graph showing that D-lactate enhances the suppression of metabolic activity in K. pneumoniae WACC790 in the presence of antibiotics. A: Ciprofloxacin. B: Colistin. Relative metabolic activity is shown in black for antibiotic treatment alone and in gray for antibiotic + A: 0.1 mM D-lactate or B: 10 mM D-lactate. [Figure 9] Figure 9 is a graph showing that D-lactate enhances the suppression of metabolic activity in S. pneumoniae D39 in the presence of antibiotics. A: Amoxicillin. B: Ampicillin. Relative metabolic activity is shown in black for antibiotic treatment alone and in gray for antibiotic + 10 mM D-lactate. [Figure 10] Figure 10 is a graph showing that D-lactate enhances the suppression of metabolic activity in S. pneumoniae D39 in the presence of antibiotics. A: Ceftriaxone. B: Tetracycline. C: Vancomycin. Relative metabolic activity is shown in black for antibiotic treatment alone and in gray for antibiotic + 10 mM D-lactate. [Figure 11] Figure 11 shows the histograms of P. aeruginosa WACC91 suspension cultures after adding D-lactic acid or L-lactic acid at 2-fold serial dilutions. [Figure 12] Figure 12 shows graphs of the total bacterial counts from the blood of control and treated mice 4 hours after infection. Significant differences between the control and treated groups were determined by an unpaired t-test (two-tailed; *=p<0.05; **=p<0.01; ns=not significant). The dashed horizontal line indicates the detection limit. [Figure 13]Figure 13 shows representative images of two CD1 male mice (ventral and dorsal) at a specified time after IP challenge with colR E. coli Xen14, demonstrating that treatment with sodium D-lactate and colistin was more effective than colistin alone. [Figure 14] Figure 14 shows a graph of the survival analysis of infected mice. Five out of eight mice in the D-sodium lactate + 1 mg / kg colistin combination group, and three out of eight mice in the 1 mg / kg colistin-treated group, survived infection by colR E. coli Xen14 until the end of the experiment (72 hours post-infection). However, all control mice and mice treated with D-sodium lactate alone succumbed to infection by 12 hours after the challenge. [Modes for carrying out the invention]

[0056] Description of the Invention Detailed description of the invention 2-hydroxycarboxylic acids, such as lactic acid, are known to be bactericidal due to their ability to lower environmental pH and disrupt cell membrane integrity. These antimicrobial properties have led to their use in food production.

[0057] However, this disclosure has surprisingly found that one enantiomer of an enantiomer of an enantiomer of a 2-hydroxycarboxylic acid, referred to in this application as “preferred enantiomer of 2-hydroxycarboxylic acid,” “preferred enantiomer,” or “preferred 2-hydroxycarboxylic acid,” is effective in dysregulation of the metabolism of planktonic bacteria. This dysregulation effect is not provided by other enantiomers of the enantiomer of the enantiomer of a 2-hydroxycarboxylic acid, also known as “unpreferred enantiomer of 2-hydroxycarboxylic acid,” “unpreferred enantiomer,” or “unpreferred 2-hydroxycarboxylic acid.” In some situations, it has been found that the unpreferred enantiomer is not only inactive in dysregulation but can also potentially prevent the preferred enantiomer from having its effect. Preferably, dysregulation of the metabolism of planktonic bacteria is suppression of the metabolism of planktonic bacteria.

[0058] The metabolic dysregulation effect is not a bactericidal effect; both enantiomers of enantiomerous 2-hydroxycarboxylic acids such as lactic acid are known to have equal bactericidal effects. In some cases, the bactericidal effect of the two enantiomers is due to their effect on pH. In this disclosure, the preferred enantiomer is used at a lower concentration than the preferred enantiomer would be used if it were used for its bactericidal effect.

[0059] The term "metabolic dysregulation" encompasses both the suppression and enhancement of bacterial vitality. Dysregulation may include changes in aerobic respiration, metabolic activity, suppression of NADH or NADPH production, changes in bacterial viability, changes in the development of resistance to antimicrobial compounds (bacteriostatic and bactericidal compounds), and / or changes in bacterial susceptibility to antimicrobial compounds (bacteriostatic and bactericidal compounds).

[0060] It has been found that the addition of a preferred enantiomer of 2-hydroxycarboxylic acid in the absence of an antimicrobial compound generally induces a suppression of the metabolism of planktonic bacteria. However, in some cases, it leads to an enhancement of metabolism. This dysregulation of bacterial metabolism has been found to cause planktonic bacteria to become more sensitive to antimicrobial compounds. Where necessary, this dysregulation of planktonic metabolism can result in bacterial sensitization to antimicrobial compounds.

[0061] In some cases, the metabolism of planktonic bacteria remains suppressed in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound. In other cases, the metabolism of planktonic bacteria may be enhanced in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound.

[0062] The metabolic dysregulation of planktonic bacteria by preferred enantiomers of 2-hydroxycarboxylic acid may be one of the following: - Inhibition of metabolism in the presence of 2-hydroxycarboxylic acid alone, and inhibition of metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds. - Inhibition of metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhancement of metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds. - Enhancement of metabolism in the presence of 2-hydroxycarboxylic acid alone, and enhancement of metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds. - Enhancement of metabolism in the presence of 2-hydroxycarboxylic acid alone, but inhibition of metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds. - No effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, however, inhibition of metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds. - No effect on metabolism in the presence of 2-hydroxycarboxylic acid alone, but enhanced metabolism in the presence of 2-hydroxycarboxylic acid and antimicrobial compounds.

[0063] The timing of exposing bacteria to 2-hydroxycarboxylic acid may be before, simultaneously with, or after the introduction of the planktonic bacteria to the antimicrobial compound.

[0064] The term "inhibition of planktonic bacterial metabolism" includes inhibition of bacterial vitality, e.g., inhibition of aerobic respiration, inhibition of mitochondrial metabolic activity, inhibition of NADH or NADPH production, inhibition of bacterial viability, inhibition of the development of resistance to antimicrobial compounds (bacteriostatic and bactericidal compounds), and / or enhancement of bacterial susceptibility to antimicrobial compounds (bacteriostatic and bactericidal compounds). Inhibition can be measured by comparing the metabolism of bacteria in the presence of a preferred enantiomer of 2-hydroxycarboxylic acid with the metabolism of the same bacteria in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid. If bacterial metabolism is low in the presence of 2-hydroxycarboxylic acid, inhibition of planktonic bacterial metabolism has occurred. The percentage change depends on the metabolic parameter used (e.g., resazurin reduction, or decrease in the MIC of the concomitant antimicrobial agent, or enhancement of in vivo antimicrobial efficacy) and may be 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100%, or any range thereof.

[0065] The term "enhancement of planktonic bacterial metabolism" includes enhancement of bacterial vitality, e.g., enhancement of aerobic respiration, enhancement of mitochondrial metabolic activity, enhancement of NADH or NADPH production, enhancement of bacterial viability, and / or enhancement of bacterial susceptibility to antimicrobial compounds (bacteriostatic and bactericidal compounds). Enhancement can be measured by comparing the metabolism of the bacteria in the presence of a preferred enantiomer of 2-hydroxycarboxylic acid with the metabolism of the same bacteria in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid. If bacterial metabolism is higher in the presence of 2-hydroxycarboxylic acid, then enhancement of planktonic bacterial metabolism has occurred. The percentage change depends on the metabolic parameter used (e.g., resazurin reduction, or decrease in the MIC of the concomitant antimicrobial agent, or enhancement of in vivo antimicrobial efficacy) and may be 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100%, or any range thereof. Enhancement of the metabolism of planktonic bacteria preferably occurs after metabolic inhibition occurs, and preferably after the addition of the antimicrobial compound. However, in some cases, enhancement of the metabolism of planktonic bacteria may occur upon exposure to a preferred enantiomer of 2-hydroxycarboxylic acid alone, without being preceded by inhibition and / or the presence of an antimicrobial compound.

[0066] This disclosure has found that preferred enantiomers of 2-hydroxycarboxylic acids, including D-lactic acid, are small, water-soluble compounds that can be used for the purpose of inhibiting the metabolism of planktonic bacteria. Planktonic bacterial infections and contaminations can be treated by preferred enantiomers of 2-hydroxycarboxylic acids alone, through their ability to disrupt bacterial metabolism. In such cases, metabolic disruption is preferably metabolic inhibition.

[0067] Alternatively, preferred enantiomers of 2-hydroxycarboxylic acid can be used in combination with other antimicrobial compounds (both bactericidal and bacteriostatic antimicrobial compounds) or therapeutic agents. The antimicrobial compounds may be antibiotic or non-antibiotic antimicrobial compounds. The dysregulatory activity of preferred enantiomers of 2-hydroxycarboxylic acid enhances the bactericidal, bacteriostatic, antibiofilm, or disinfectant efficacy of the antimicrobial compound or antimicrobial therapy. In such cases, metabolic disruption may be metabolic inhibition or metabolic enhancement. For example, preferred enantiomers of 2-hydroxycarboxylic acid may induce metabolic inhibition, followed by metabolic enhancement upon introduction of an antimicrobial compound. Alternatively, preferred enantiomers of 2-hydroxycarboxylic acid may induce metabolic inhibition, which is maintained in the presence of an antimicrobial compound. However, in some cases, enhancement of the metabolism of planktonic bacteria may occur upon exposure to preferred enantiomers of 2-hydroxycarboxylic acid alone, without being preceded by inhibition and / or the presence of an antimicrobial compound. In such cases, metabolic suppression may follow metabolic enhancement in the presence of 2-hydroxycarboxylic acids and antimicrobial compounds.

[0068] In this disclosure, preferred enantiomers of 2-hydroxycarboxylic acids may act as “antibiotic resistance breakers” (ARBs). ARBs are compounds administered with antibiotics to overcome bacterial resistance to those antibiotics by improving the antibiotic’s potency or by inhibiting bacterial resistance mechanisms. When preferred enantiomers of 2-hydroxycarboxylic acids act as ARBs, they may improve the potency of the antibiotic by lowering its minimum inhibitory concentration (MIC). For example, preferred enantiomers of 2-hydroxycarboxylic acids may lower the MIC of an antibiotic to a value lower than the MIC of the antibiotic when administered as monotherapy.

[0069] While not bound by any particular theory, it is thought that metabolic dysregulation induced by preferred enantiomers of 2-hydroxycarboxylic acids may allow 2-hydroxycarboxylic acids to act as ARBs, improving bacterial susceptibility to antibiotics and / or increasing antibiotic potency.

[0070] If the metabolic dysregulation induced by a preferred enantiomer of 2-hydroxycarboxylic acid is metabolic suppression, the effect of 2-hydroxycarboxylic acid acting as an ARB may be reduced efflux, the inability of bacteria to modify the antibiotic after it has been taken up into the bacteria, or an increase in the amount of antibiotic retained within the bacterial cell through another mechanism.

[0071] When bacteria are in the presence of both a preferred enantiomer and an antimicrobial compound of 2-hydroxycarboxylic acid, if the metabolism of planktonic bacteria is enhanced, the effect of 2-hydroxycarboxylic acid acting as an ARB may be increased uptake of the antimicrobial compound or increased free radical generation.

[0072] As used herein, the term “planktonic bacterium” refers to bacteria that are free-swimming or free-flowing and not located within a biofilm. Although bacteria are free-swimming or free-flowing, the swimming or flow may be on a solid surface. Therefore, planktonic bacteria can be present on surfaces. Nevertheless, their presence on a surface is not within a biofilm or bound to a biofilm; planktonic bacteria are not attached to a surface. At the micrometer level, the planktonic habitats of prokaryotes may also include water films around soil particles, saliva in the mouth, fluids in the intestinal lumen, serum in blood vessels, urine in the bladder and urinary tract, aqueous compositions flowing through the xylem and / or sieve of plants, surface water on plant leaves, and within soil substrates. Planktonic bacteria can eventually grow into biofilms by settling on surfaces, attaching to surfaces, and then pushing out the extracellular matrix. However, the bacteria targeted by this disclosure are bacteria that have not yet developed the characteristics of a biofilm, or individual bacteria that have been released from a biofilm and become free-flowing, capable of settling elsewhere and restarting the biofilm process.

[0073] The planktonic bacteria described herein may be bacteria that are unable to develop into biofilms and survive in a planktonic form.

[0074] The planktonic bacteria of this disclosure may be bacteria that are released spontaneously or induced from a biofilm-binding colony, for example, due to the disruption of the biofilm matrix.

[0075] Compared to bacteria within biofilms, planktonic bacteria often possess upregulated DNA repair genes (such as bacterial SOS response genes). Bacteria released from biofilms and becoming planktonic may retain some or all of the characteristics of the biofilm-associated "parent" population, and have therefore been further reported to exhibit increased antibiotic resistance. These released planktonic populations are difficult to remove from situations requiring sterility and often lead to recurrent infections.

[0076] composition This disclosure provides a composition for dysregulating the metabolism of planktonic bacteria, the composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid such as D-lactic acid.

[0077] This disclosure further provides compositions for dysregulating the metabolism of planktonic bacteria, the compositions comprising preferred enantiomers of 2-hydroxycarboxylic acids and antimicrobial compounds.

[0078] The disclosure further provides a composition for sensitizing planktonic bacteria to an antimicrobial compound. The composition comprises a preferred enantiomer of 2-hydroxycarboxylic acid, which induces metabolic dysregulation in planktonic bacteria.

[0079] The disclosure further provides a composition for sensitizing planktonic bacteria to an antimicrobial compound, the composition comprising a preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, the preferred enantiomer of 2-hydroxycarboxylic acid inducing metabolic dysregulation in planktonic bacteria.

[0080] Dysregulation of bacterial metabolism can lead to sensitization. Sensitization may be due to an enhancing or synergistic effect. In one aspect, dysregulation of the metabolism of planktonic bacteria is suppression of planktonic metabolism; in another aspect, it is enhancement of planktonic metabolism; and in yet another aspect, it is either suppression or enhancement followed by the other.

[0081] 2-hydroxycarboxylic acid The 2-hydroxycarboxylic acid of the present invention has stereochemistry, and one enantiomer of its stereochemistry is preferred. The 2-hydroxycarboxylic acid of the present disclosure having preferred stereochemistry is referred to herein as “preferred enantiomer of 2-hydroxycarboxylic acid,” “preferred enantiomer,” or “preferred 2-hydroxycarboxylic acid.” The 2-hydroxycarboxylic acid is optionally a hydrophilic 2-hydroxycarboxylic acid. In one embodiment, the preferred enantiomer of the 2-hydroxycarboxylic acid is D-lactic acid. The preferred 2-hydroxycarboxylic acid of the present invention may be a salt of the preferred 2-hydroxycarboxylic acid, and may optionally be a pharmaceutically acceptable salt.

[0082] If the preferred 2-hydroxycarboxylic acid is not D-lactic acid, the preferred 2-hydroxycarboxylic acid used in this disclosure may have a three-dimensional orientation of the hydroxyl group such that the hydroxyl group of the 2-hydroxycarboxylic acid corresponds to the absolute stereochemistry of the chiral center of the corresponding D-lactic acid, as shown below. [ka]

[0083] The preferred 2-hydroxycarboxylic acids of the above general formula are those in which the substituent R is hydrogen, halogen (F, Cl, Br, I), methyl, ethyl, n-propyl, isopropyl, cyclopropyl, n-butyl, iso-butyl, sec-butyl, tert-butyl, cyclobutyl, n-pentyl, isopentyl, neopentyl, cyclopentyl, n-hexyl, 2-methylpentyl, 3-methylpentyl, 2,3-dimethylbutyl, 2,2-dimethylbutyl, cyclohexyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 3-ethylpentyl, 2,2,3-trimethylbutyl, cycloheptyl, phenyl, benzyl, furanyl, tetrahydrofuranyl, ethenyl, vinyl, or allyl These substituents may be selected from the group consisting of clotyl, isopentenyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 2-methylpropa-2-enyl, hexenyl, butadienyl, pentadienyl, hexadienyl, 2,3-dimethyl-2-butenyl, heptenyl, octenyl, octatrienyl, butadienyl, 1,3-pentadienyl, 1,4-pentadienyl, cyclopentadienyl, and cyclohexadienyl, and each of these substituents (except hydrogen and halogens) may be unsubstituted or substituted with one or more substituents selected from the group consisting of hydrogen, halogens, hydroxyl, methoxyl, ethoxyl, propoxyl, butoxyl, carboxylic acids, amide or ester substituents, and pharmaceutically acceptable salts thereof.

[0084] The preferred 2-hydroxycarboxylic acid base addition salts of the present invention can be prepared from inorganic and organic bases. Examples of salts derived from inorganic bases include sodium salts, potassium salts, lithium salts, ammonium salts, calcium salts, magnesium salts, copper salts, zinc salts, gallium salts, and silver salts. Examples of salts derived from organic bases include, but are not limited to, salts of primary, secondary, and tertiary amines, such as alkylamines, dialkylamines, trialkylamines, substituted alkylamines, di(substituted alkyl)amines, tri(substituted alkyl)amines, alkenylamines, dialkenylamines, trialkenylamines, substituted alkenylamines, di(substituted alkenyl)amines, tri(substituted alkenyl)amines, cycloalkylamines, di(cycloalkyl)amines, tri(cycloalkyl)amines, substituted cycloalkylamines, disubstituted cycloalkylamines, trisubstituted cycloalkylamines, cycloalkenylamines, di(cycloalkenyl) Examples include (1)amines, tri(cycloalkenyl)amines, substituted cycloalkenylamines, disubstituted cycloalkenylamines, trisubstituted cycloalkenylamines, arylamines, diarylamines, triarylamines, heteroarylamines, diheteroarylamines, triheteroarylamines, heterocyclic amines, diheterocyclic amines, triheterocyclic amines, and mixed di- and tri-amines selected from the group consisting of alkyl, substituted alkyl, alkenyl, substituted alkenyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, heteroaryl, heterocyclic, etc., in which at least two substituents on the amine are different. Amines in which two or three substituents combine with an amino nitrogen to form a heterocyclic or heteroaryl group are also included.

[0085] The preferred 2-hydroxycarboxylic acid acid addition salts of the present invention can be prepared from inorganic and organic acids. Examples of inorganic acids that can be used include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid. Examples of organic acids that can be used include acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid.

[0086] Examples of acceptable salts include iodide, acetate, phenylacetate, trifluoroacetate, acrylate, ascorbate, benzoate, chlorobenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, methylbenzoate, o-acetoxybenzoate, naphthalene-2-benzoate, bromide, isobutyrate, phenylbutyrate, γ-hydroxybutyrate, β-hydroxybutyrate, buty-1,4-dioete, hexyn-1,4-dioete, hexyn-1,6-dioete, caproate, caprylate, chloride, cinnamate, citrate, decanoate, formate, fumarate, glycolate, heptanoate, hiplate, lactate, maleate, maleate, hydroxymaleate, malonate, mandelate, mesiolate. Examples include phosphates, nicotinates, isonicotinates, nitrates, oxalates, phthalates, terephthalates, phosphates, monohydrogen phosphates, dihydrogen phosphates, metaphosphates, pyrophosphates, propiolates, propionates, phenylpropionates, salicylates, sebacates, succinates, sverates, sulfates, bisulfates, pyrosulfates, sulfites, bisulfites, sulfonates, benzenesulfonates, p-bromophenylsulfonates, chlorobenzenesulfonates, propanesulfonates, ethanesulfonates, 2-hydroxyethanesulfonates, methanesulfonates, naphthalene-1-sulfonates, naphthalene-2-sulfonates, p-toluenesulfonates, xylenesulfonates, and tartarates.

[0087] The compositions of this disclosure may contain 2-hydroxycarboxylic acids with undesirable stereochemistry, such as L-lactic acid. The 2-hydroxycarboxylic acids of this disclosure having undesirable stereochemistry are referred to herein as “undesirable enantiomers of 2-hydroxycarboxylic acids,” “undesirable enantiomers,” or “undesirable 2-hydroxycarboxylic acids.” The percentage of undesirable 2-hydroxycarboxylic acids may be less than 20%. Alternatively, the compositions of this disclosure may not contain undesirable 2-hydroxycarboxylic acids. Therefore, the compositions may contain only 2-hydroxycarboxylic acids with preferred stereochemistry.

[0088] If the undesirable 2-hydroxycarboxylic acid is not L-lactic acid, the undesirable 2-hydroxycarboxylic acid according to this disclosure is one in which, with respect to the three-dimensional orientation of the hydroxyl group, the hydroxyl group of the 2-hydroxycarboxylic acid corresponds to the absolute stereochemistry of the chiral center of the corresponding L-lactic acid, and the absolute stereochemistry of the chiral center is as shown below: [ka]

[0089] Generally, most lactic acid bacteria primarily produce L-lactic acid. While not bound by any theory, it is thought that many bacteria have evolved to avoid or denature L-lactic acid to prevent the effects of organic acids on the bacterial membrane. Again, while not bound by any theory, bacteria have a low ability to avoid or denature the D-isomer of lactic acid, and as a result, the D-isomer may have a metabolic disruptive effect.

[0090] Highly pure L-lactic acid can be produced by a wide range of microorganisms, including bacteria, fungi, algae, and cyanobacteria. On the other hand, most D-lactic acid-producing microorganisms produce either a racemic mixture or other organic acids such as acetic acid or succinic acid (Alexandri et al. (2019) Food Tech & Biotech 57(3):293-304).

[0091] The compositions of this disclosure may contain 0.001% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid, such as D-lactic acid. The compositions may contain 0.001%, 0.01%, 0.1%, 1%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient, and / or other activators. For example, an ophthalmic composition may contain 0.001% to 1% of a preferred enantiomer of a 2-hydroxycarboxylic acid. Washing and immersion, bandage and wipe, topical, inhalation, or oral compositions may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid.

[0092] The compositions of this disclosure may contain undesirable enantiomers of 2-hydroxycarboxylic acid, the percentage of which is less than 50%, less than 40%, less than 30%, less than 20%, less than 10%, less than 5%, or less than 1%. For example, the percentage of undesirable enantiomers of 2-hydroxycarboxylic acid may be less than 20%, less than 18%, less than 16%, less than 14%, less than 12%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%. The percentage of undesirable enantiomers of 2-hydroxycarboxylic acid may be less than 20%.

[0093] Alternatively, the compositions of the present disclosure may not contain any undesirable enantiomers of 2-hydroxycarboxylic acid. Therefore, the compositions may contain only the preferred enantiomer of 2-hydroxycarboxylic acid as 2-hydroxycarboxylic acid. For example, they may contain only D-lactic acid as lactic acid. It has been found that compositions containing an enantiomerically pure amount of the preferred enantiomer of 2-hydroxycarboxylic acid have a metabolic dysregulatory effect on planktonic bacteria, compositions containing an enantiomerically pure amount of the undesirable enantiomer of 2-hydroxycarboxylic acid have little to no metabolic dysregulatory effect, and compositions containing a 1:1 ratio of the preferred enantiomer of 2-hydroxycarboxylic acid to the undesirable enantiomer of 2-hydroxycarboxylic acid have a reduced metabolic dysregulatory effect or no metabolic dysregulatory effect on planktonic bacteria compared to administering the pure preferred enantiomer of 2-hydroxycarboxylic acid.

[0094] The compositions of this disclosure may optionally contain an enantiomerized amount of a preferred enantiomer of a 2-hydroxycarboxylic acid. The compositions may also contain an enantiomerized pure preferred enantiomer of a 2-hydroxycarboxylic acid.

[0095] If necessary, the concentration of the preferred enantiomer of 2-hydroxycarboxylic acid used in the composition of the present disclosure is lower than the concentration of the preferred enantiomer of 2-hydroxycarboxylic acid required to kill bacteria.

[0096] Without being bound by any theory, this disclosure assumes the need for a 2-hydroxycarboxylic acid having a chiral center at position 2. It is understood that the chiral center is at position 2 if the aliphatic chain has a length of at least 3 carbon atoms. By this definition, glycolic acid [2-hydroxyacetic acid] is not chiral because its length is less than 3 carbon atoms.

[0097] 2-hydroxycarboxylic acids may also be hydrophilic 2-hydroxycarboxylic acids; that is, 2-hydroxycarboxylic acids can be soluble in water if necessary. While not bound by any theory, the water-soluble nature of 2-hydroxycarboxylic acids is thought to contribute to their dysregulation of the membranes and metabolism of planktonic bacteria.

[0098] In general, 2-hydroxycarboxylic acids having seven or fewer carbon atoms (including the carbon atoms of the hydroxycarboxylic acid moiety) have been found to be more water-soluble. Therefore, as needed, the 2-hydroxycarboxylic acids of this disclosure may contain seven or fewer carbon atoms. For example, 2-hydroxypropanoic acid (also known as lactic acid) contains three carbon atoms, 2-hydroxypentanoic acid (also known as 2-hydroxyvaleric acid) contains five carbon atoms, and 2-hydroxycaproic acid (also known as 2-hydroxyhexanoic acid) contains six carbon atoms. For example, the 2-hydroxycarboxylic acids of this disclosure may contain six or fewer carbon atoms. In one example, the 2-hydroxycarboxylic acids of this disclosure may contain five or fewer carbon atoms.

[0099] If necessary, the molecular weight of the 2-hydroxycarboxylic acid is less than 800, less than 700, less than 600, or less than 500. For example, the molecular weight of the 2-hydroxycarboxylic acid is less than 200, less than 180, less than 150, or less than 100. For example,

[0100] If necessary, the absolute water solubility of 2-hydroxycarboxylic acid is greater than approximately 0.5 g / L, 0.75 g / L, or 1.0 g / L at pH approximately 5-10, pH approximately 9-6, or pH approximately 6-8.

[0101] If necessary, the absolute water solubility of 2-hydroxycarboxylic acid exceeds approximately 0.5 g / L, 0.75 g / L, or 1.0 g / L at a temperature of approximately 20°C.

[0102] The water solubility of a 2-hydroxycarboxylic acid can alternatively be defined by its LogP (i.e., its octanol-water partitioning efficiency). LogP is equal to LogKow Also known as the n-octanol-water partition ratio. In principle, LogP:octanol-water < 1.0 = water-soluble; LogP:octanol-water > 1.0 = hydrophobic. Where necessary, the 2-hydroxycarboxylic acids of this disclosure have LogP:octanol-water < 1.0. Some compounds having LogP: < 1.0 may have more than 7 carbon atoms; however, their structure allows for water solubility despite the additional carbon atoms. For example, 2-hydroxy-2-phenylacetic acid (also known as mandelic acid) has 8 carbon atoms, but due to its ring structure, it has a LogP of 0.67 and is therefore water-soluble.

[0103] Some 2-hydroxycarboxylic acids with special structures, such as 2-hydroxy-2-phenylacetic acid (also known as mandelic acid), which has a six-membered ring and a total of eight carbon atoms, can still be water-soluble and may have a LogP:octanol-water < 1.0. Such 2-hydroxycarboxylic acids are considered suitable for the present disclosure.

[0104] 2-hydroxycarboxylic acids may have substituents such as hydrogen, halogen, hydroxyl, methoxyl, ethoxyl, propoxyl, butoxyl, carboxylic acid, amide, or ester substituents.

[0105] 2-hydroxycarboxylic acids can be pharmaceutically acceptable. For example, 2-hydroxycarboxylic acids are GRAS ("generally recognized as safe") as defined in Sections 201(s) and 409 of the U.S. Food, Drug, and Cosmetic Act or equivalent regulations.

[0106] The 2-hydroxycarboxylic acid can be selected from the group of acids, esters, salts, amides, or other derivatives, consisting of lactic acid, glycolic acid (2-hydroxyacetic acid), tartaric acid (2,3-dihydroxysuccinic acid), mandelic acid, 1-hydroxycyclohexane-1-carboxylic acid, 2-hydroxy-2-(tetrahydrofuran-2-yl)acetic acid, 2-hydroxy-2-(2-furanyl)ethaneic acid, 2-hydroxy-2-phenylpropionic acid, 2-hydroxy-2-methylpropionic acid, 2-hydroxy-2-methylbutanoic acid, 2-hydroxybutanoic acid, 2-hydroxypentanoic acid, or mixtures thereof.

[0107] The 2-hydroxycarboxylic acid may be selected from a list including 2-hydroxypropanoic acid (also known as lactic acid), 2-hydroxypentanoic acid (also known as 2-hydroxyvaleric acid), 2-hydroxybutyric acid, 2-hydroxyacetic acid (also known as glycolic acid), 2-hydroxyhexanoic acid (also known as 2-hydroxycaproic acid), and 2-hydroxy-2-phenylacetic acid (also known as mandelic acid).

[0108] The 2-hydroxycarboxylic acid may be selected from a list including D-lactic acid, 2-hydroxypentanoic acid, 2-hydroxybutyric acid, 2-hydroxyhexanoic acid, (2)-hydroxyphenylacetic acid, and 2-hydroxy-2-phenylacetic acid. Preferred 2-hydroxycarboxylic acids are those listed in Table 1 and / or Table 2. [Table 1] [Table 2]

[0109] If necessary, the 2-hydroxycarboxylic acid has both fewer than 7 carbon atoms and LogP: octanol-water < 1.0.

[0110] If necessary, the 2-hydroxycarboxylic acid of this disclosure is lactic acid that asheses two enantiomers. If necessary, preferred enantiomers of the 2-hydroxycarboxylic acid are (2R)-2-hydroxypropanoic acid and D-lactic acid, also known as (R)-lactic acid. [ka]

[0111] Preferred enantiomers of 2-hydroxycarboxylic acids can be synthesized via cell-free pathways (e.g., enzyme-catalyzed reactions). They can be derived from bacteria such as lactic acid bacteria, e.g., Lactobacillus or Bifidobacterium, from supernatants derived from cultured bacteria such as multi-strain Gram-positive and / or Gram-negative probiotic bacterial strains, from natural sources such as 2-hydroxy-2-phenylacetic acid (mandelic acid) derived from almonds, olive oil, and beer, or from other starchy biomass.

[0112] If the preferred enantiomer of 2-hydroxycarboxylic acid is D-lactic acid, D-lactic acid can be synthesized from Lactobacillus or from the supernatant of cultured bacteria such as polystrain Gram-negative probiotic bacterial strains via a cell-free pathway (e.g., enzyme-catalyzed reaction).

[0113] The D-lactic acid in this disclosure is preferably a monomer. Therefore, the D-lactic acid is, if necessary, not polylactic acid (also known as PLA, poly-D-lactic acid [PDLA], or poly-L-lactic acid [PLLA]) or a similar polymer. PLA is a polymer obtained by ring-opening polymerization of monomer lactide (a cyclic dimer of lactic acid).

[0114] antimicrobial compounds The methods of the present disclosure further provide preferred enantiomers of 2-hydroxycarboxylic acid for co-administration in combination with an antimicrobial compound, the preferred enantiomer of 2-hydroxycarboxylic acid increasing the activity of the antimicrobial compound against planktonic bacteria. The preferred enantiomer of 2-hydroxycarboxylic acid may have the same composition as the antimicrobial compound or a different composition. Planktonic bacteria may cause infection in a subject, infection within or on a plant, infection in the soil on which the plant is growing, or be present on a surface such as a non-living surface. Planktonic bacteria may be free-flowing planktonic bacteria in a liquid or gas.

[0115] The term "antibiotic compound" refers to a compound that can kill bacteria or reduce their vitality. Antimicrobial compounds may be antibiotics or agents with different modes of antimicrobial action, such as bacteriophages, disinfectants, salines, chlorine compounds (such as bleaches), iodine compounds, copper compounds, or heavy metals. This term includes bactericidal, bacteriostatic, antibiofilm, or disinfectant antimicrobial compounds. Antimicrobial compounds may also be antibiofilm compounds.

[0116] The term "antibiotic" refers to an antimicrobial compound that is active against bacteria and used in a medical or veterinary setting for prevention or treatment. They may either kill bacteria (bactericidal) or inhibit bacterial growth (bacteriostatic). Antibiotics may be of natural origin (compounds derived from screening bacteria and fungi for antibiotic compounds, e.g., penicillin) or of laboratory origin (compounds derived from chemically determining targets for antibiotic compounds, e.g., sulfamethoxazole). Antibiotics may be derivatives of naturally derived or laboratory-derived compounds.

[0117] If the antimicrobial compound administered with the preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure is an antibiotic, it may be selected from one or more of the following classes. [Table 6]

[0118] The term "sensitization" refers to an increase in the effectiveness of an antimicrobial compound against planktonic bacteria, for example, by reducing the minimum inhibitory concentration (MIC) of the antimicrobial compound. The MIC is the lowest concentration of an antibiotic at which visible bacterial growth is inhibited. Sensitization can occur due to an enhancing or synergistic effect.

[0119] Preferably, the planktonic bacteria are resistant to one or more antimicrobial compounds.

[0120] As used herein, the term “synergistic effect” refers to a case where 2-hydroxycarboxylic acid and an antimicrobial compound, such as an antibiotic, act together to produce a more potent effect than when each active substance is applied individually. For example, in this disclosure, a synergistic effect may occur if the combined effect of 2-hydroxycarboxylic acid and an antimicrobial compound on planktonic bacteria is greater than the sum of their effects when applied separately. A synergistic effect may occur if planktonic bacteria are susceptible to the antimicrobial compound (e.g., by increasing the effect of the antimicrobial compound so that the amount of antimicrobial compound used to obtain the same effect can be reduced, by lowering the MIC, etc.), or if planktonic bacteria are resistant to the antimicrobial compound (e.g., by increasing the ability of the antimicrobial compound to act on planktonic bacteria to such an extent that the antimicrobial compound can be effectively used even if it was previously ineffective due to resistance, by lowering the MIC to an effective value that can be safely, easily, and cost-effectively administered, etc.).

[0121] The measurement of the synergistic effect in the combination of a preferred enantiomer of a 2-hydroxycarboxylic acid and an antimicrobial compound can be achieved using a simple plate assay in which the compounds are applied to separate paper discs and diffused on an agar plate inoculated with the target bacteria. The synergistic effect is indicated by enhanced killing at the interface of the diffusion zones. The quantitative measurement of the synergistic effect can be measured using a simple checkerboard strategy, systematically diluting the concentrations of both agents to identify the concentrations of both agents that achieve the most potent interaction.

[0122] The synergistic effect is often measured by calculation of the fractional inhibitory concentration index (FICI): FICI = (MIC AB / MIC A ) + (MIC BA / MICM B ) MIC AB is the minimum inhibitory concentration (MIC) of drug A tested in combination, MIC A is the MIC of drug A tested alone, MIC BA is the MIC of drug B tested in combination, and MIC B is the MIC of drug B tested alone. An FICI of 0.50 or less is defined as a synergistic effect (i.e., a four-fold decrease in MIC), an FICI between 0.50 and 1.00 represents an additive effect, and an FICI between 1.00 and 4.00 is defined as no interaction. An FICI greater than 4.00 represents an antagonistic effect.

[0123] However, since 2-hydroxycarboxylic acid itself does not possess antimicrobial effects and rather merely enhances the effects of antimicrobial compounds, it can be difficult to determine the FICI value of a preferred enantiomer of 2-hydroxycarboxylic acid and a combination of antimicrobial compounds. In other cases, the antimicrobial compound may not have an MIC in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid. If neither the preferred enantiomer of 2-hydroxycarboxylic acid nor the antimicrobial compound has a measurable MIC, a concentration of 2-hydroxycarboxylic acid can be used to enable the determination of the MIC for the antimicrobial compound. In one example, the concentration of 2-hydroxycarboxylic acid that reduces the MIC of the antimicrobial compound can be used as an indicator of potency. For example, a reduction of the MIC to one-quarter can be used as an indicator of potency (i.e., a reduction of the MIC to one-quarter). More importantly, the concentration of the preferred enantiomer of 2-hydroxycarboxylic acid that reduces the MIC of the antimicrobial compound in resistant bacteria to below or equal to the breakpoint concentration, i.e., the concentration of the antimicrobial compound that defines whether the strain is resistant or susceptible. Therefore, the rescue concentration (RC) is defined as the concentration of 2-hydroxycarboxylic acid required to reduce the effective concentration of the antimicrobial compound to below or equal to the breakpoint (Wright et al. (2016) Trends Microbiol. 24(11):862-871).

[0124] As used herein, the term “enhancing effect” refers to the ability of 2-hydroxycarboxylic acid to increase the activity of antimicrobial compounds such as antibiotics. For example, in this disclosure, an enhancing effect may occur when planktonic bacteria are resistant to an antimicrobial compound (and therefore the antimicrobial compound alone is ineffective, i.e., has no MIC). However, when an antimicrobial compound is combined with a 2-hydroxycarboxylic acid, the response of planktonic bacteria to the antimicrobial compound is enhanced (i.e., the antimicrobial compound has a more significant effect on the bacteria, either bacteriostatic or bactericidal, i.e., its MIC is now determinable).

[0125] Preferred enantiomers of 2-hydroxycarboxylic acids of this disclosure have the ability to combat bacterial resistance through multiple mechanisms, including enhancing the uptake of antimicrobial compounds, inhibiting the leaching of antimicrobial compounds, disrupting signaling pathways, preventing modification of antimicrobial compounds, and / or preventing modification of antimicrobial compound target sites.

[0126] If the dysregulation of bacterial metabolism is due to the suppression of metabolism by a preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure, the metabolic suppression may act to retain the antimicrobial compound in the planktonic bacteria, for example, by slowing the efflux of the antimicrobial compound as metabolism (including efflux pumps) is suppressed. Alternatively, the antimicrobial compound may be retained within the planktonic bacteria by slowing the rate at which the antimicrobial compound is modified in conjunction with the suppression of metabolism (including enzyme production and activity).

[0127] If the dysregulation of bacterial metabolism is due to metabolic enhancement by a preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure, the metabolic enhancement may act, for example, by increasing the uptake of antimicrobial compounds. Alternatively, the metabolic enhancement may result in increased free radical production, making bacterial cells more sensitive to antimicrobial compounds.

[0128] If necessary, the pH of the compositions of this disclosure is about 6.5 to 8.0, for example, about 7.0 and 7.4. It has been previously found that bacteria become more resistant to antimicrobial therapy as the pH decreases. The preferred pH helps to avoid bacterial resistance to the effects of the preferred enantiomer of 2-hydroxycarboxylic acid. Buffers may be added to adjust the pH level of the compositions. If necessary, the compositions of this disclosure contain tris(hydroxymethyl)aminomethane (TRIS, also known as THAM or tromethamine) or phosphate-buffered saline (PBS) as buffers. The effectiveness of the preferred enantiomer of 2-hydroxycarboxylic acid at a pH of about 6.5 to 8.0 is a further indicator that the effect on bacterial metabolism is a result of the activity of 2-hydroxycarboxylic acid itself against metabolic dysregulation, as well as the effect of low pH on killing bacteria.

[0129] While not bound by any theory, preferred enantiomers of the 2-hydroxycarboxylic acids of this disclosure are thought to work by dysregulating the metabolic rates of planktonic bacteria. Dysregulation may be inhibition or enhancement, or one of the two, followed by the other.

[0130] Metabolic inhibition can lead to increased susceptibility of planktonic bacteria to antimicrobial compounds. Unlike the activity of any particular antimicrobial compound, metabolic inhibition can occur in the absence of any further antimicrobial compounds.

[0131] Metabolic enhancement can result in increased susceptibility of planktonic bacteria to antimicrobial compounds. Unlike the activity of any antimicrobial compound, metabolic enhancement can occur in the absence of any further antimicrobial compounds. Nevertheless, it generally occurs in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound. However, in some cases, metabolic enhancement of planktonic bacteria can occur upon exposure to the preferred enantiomer of 2-hydroxycarboxylic acid alone, without being preceded by inhibition and / or the presence of an antimicrobial compound. In such cases, metabolic inhibition may follow the metabolic enhancement in the presence of both 2-hydroxycarboxylic acid and the antimicrobial compound.

[0132] If the metabolism of planktonic bacteria is dysregulated, the bacteria may be affected by one or more of the following effects: - High vulnerability of bacteria to host immune system agents, other microorganisms, or other antimicrobial compounds, including but not limited to antibiotics and bacteriophages; - A decrease in the ability of bacteria to develop resistance to antimicrobial compounds (whether external or host-derived); - Decreased cytoplasmic membrane potential or decreased cytoplasmic membrane integrity; and / or - Reduced bacterial growth.

[0133] Dysregulation of bacterial metabolism can be measured as a decrease in the MIC of an antimicrobial compound (enhanced potency), a shift in respiratory metabolism (measured by resazurin reduction), or any other measure of bacterial metabolism. All measurements can be performed either in vitro or in vivo. The degree of metabolic dysregulation should preferably be quantifiable in a clinical setting, and the degree of metabolic dysregulation should be clinically significant. Dysregulation of bacterial metabolism should result in bacterial sensitization to an antimicrobial compound and / or a decrease in the MIC of the antimicrobial compound, regardless of how the dysregulation is measured.

[0134] If necessary, the treatment regimens of this disclosure induce metabolic dysregulation of bacteria in the form of at least twofold metabolic suppression, as determined, for example, by the reduction of resazurin to resorphine. Alternatively, metabolic dysregulation of planktonic bacteria in the form of metabolic suppression can be measured via sensitization to an antimicrobial agent, which is a decrease of at least 20%, 30%, or 50% in the MIC or bMIC concentration of the antimicrobial agent.

[0135] If necessary, the treatment regimens of this disclosure may induce dysregulation of bacterial metabolism in the form of at least twofold metabolic enhancement, as determined, for example, by the reduction of resazurin to resorphine. Alternatively, metabolic dysregulation of planktonic bacteria in the form of metabolic enhancement can be measured via sensitization to an antimicrobial compound, which is a decrease of at least 20%, 30%, or 50% in the MIC or bMIC concentration of the antimicrobial compound.

[0136] Planktonic bacteria Planktonic bacteria can be Gram-positive or Gram-negative. For example, planktonic bacteria may belong to the Enterobacteriaceae family, or to the following genera: Bacillus species, Clostridium species, Campylobacter species, Pseudomonas species, Streptococcus species, Actinobacillus species, Staphylococcus species, Escherichia species, Acinetobacter species, Klebsiella species, Aeromonas species, Enterococcus species, Legionella species, and Salmonella species, Shigella species, Gardner species. The following species may be selected: ella, Haemophilus, Helicobacter, Moraxella, Mycobacterium, Neisseria, Anaerococcus, Atopobium, Bacteroides, Leptotrichia, Mobiluncus, Peptostreptococcus, Prevotella, Sneathia, Sphingomonas, Nitrospira, Mycobacterium, or Hyphomicrobium and Vibrio. The planktonic bacteria may be cyanobacteria or nontuberculous mycobacteria (NTM).

[0137] Planktonic bacteria include Pseudomonas aeruginosa, Streptococcus pneumoniae, Streptococcus mutans, Actinobacillus pleuropneumoniae, Staphylococcus epidermidis, Escherichia coli, Staphylococcus aureus, Acinetobacter baumannii, Aeromonas hydrophila, Aeromonas salmonicida, Clostridium difficile, and Enterococcus. faecium, Gardnerella vaginalis, Haemophilus influenzae, Helicobacter pylori, Moraxella bovis, Moraxella catarrhalis, Mycobacterium avium, Mycobacterium intracellulare; Mycobacterium tuberculosis, Neisseria gonorrhoeae, Neisseria meningitidis, Streptococcus pyogenes, and Vibrio cholerae.

[0138] For example, the genera Bacillus, Clostridium, and Pseudomonas are commonly found in gas and oil in pipelines. The genera Sphingomonas, Nitrospira, Legionella, Mycobacterium, or Hyphomicrobium are commonly found in water in water pipes and in air in air conditioning units. Eubacteria, including cyanobacteria, may also be present in seawater and, upon attachment, can form the basis for biofilm contamination on ship hulls.

[0139] Bacterial vaginitis may be caused by or associated with Gardnerella vaginalis and other taxa, such as fungi of the genera Anaerococcus, Atopobium, Bacteroides, Leptotrichia, Mobiluncus, Peptostreptococcus, Prevotella, Sneathia, and Clostridia, and may be treated with the metabolically dysregulated compositions of this disclosure.

[0140] Planktonic bacteria can be antibiotic-resistant strains of the bacteria listed above. For example, planktonic bacteria could be antibiotic-resistant Pseudomonas aeruginosa or antibiotic-resistant Staphylococcus aureus (e.g., MRSA).

[0141] Airborne diarrheal bacteria are typically transmitted through the "oral-fecal route," i.e., via contaminated drinking water, ice in cocktails (or soft drinks), or inadequate handwashing.

[0142] Airborne bacteria may be associated with symptoms or diseases such as sepsis, cystic fibrosis-associated pneumonia, periodontitis, diabetic ulcers and other wound infections, otitis media, sinusitis, infectious endocarditis, eye infections, serous otitis media, dysentery, respiratory infections, sexually transmitted diseases (STDs), bacterial vaginosis, and osteomyelitis.

[0143] Alternatively, planktonic bacteria may be associated with industrial and non-biological surfaces such as hydroelectric turbines, oil and gas pipelines, air conditioning units, soap dispensers, industrial water systems, and ship hulls.

[0144] In further examples, airborne bacteria can be associated with medical devices and instruments such as bags and indwelling medical devices used to preserve donated blood.

[0145] Planktonic bacteria can also be involved in the production and storage of food. For example, planktonic bacteria can be present on food preparation surfaces, storage containers, water used to wash uncooked foods (e.g., salads), or in the food itself.

[0146] Planktonic bacteria may also be associated with materials and equipment related to animal housing or livestock farming.

[0147] As used herein, “to treat” or “treatment” means to inhibit a disease or symptom, i.e., to stop or reduce its onset or at least one clinical or subclinical symptom. “To treat” or “therapy” further means to alleviate a disease or symptom, i.e., to cause regression of at least one clinical or subclinical symptom of the disease or symptom. The benefit to the treated subject is statistically significant or at least perceptible to the subject and / or the physician.

[0148] The ability of preferred enantiomers of 2-hydroxycarboxylic acids to dysregulate the metabolism of planktonic bacteria can be evaluated or measured using resazurin (7-hydroxy-3H-phenoxazine-3-one 10 oxide). Bacteria undergoing aerobic respiration have reducing cytoplasm, and under these conditions, resazurin is irreversibly reduced to a highly fluorescent resorphine. Thus, the number of “relative fluorescence units” (RFUs) is a readout of aerobic respiration / oxidative phosphorylation. This disclosure has found that, in some embodiments, administration of preferred enantiomers of 2-hydroxycarboxylic acids to planktonic bacteria results in a significant change (decrease or increase) in the rate of resazurin reduction, and therefore in a significant change (decrease or increase) in the rate of aerobic respiration / oxidative phosphorylation of the bacteria. The assays used in this disclosure determine the minimum inhibitory concentration (MIC) of the preferred enantiomer of 2-hydroxycarboxylic acid (rather than determining dose / response).

[0149] If necessary, metabolic dysregulation induced by preferred enantiomers of 2-hydroxycarboxylic acid does not result in the death of planktonic bacteria. We note that it has been suggested that administration of bactericidal compounds may result in increased metabolic rates and increased free radical production (oxidative stress leads to cell death). In consideration of this, the preferred enantiomers of 2-hydroxycarboxylic acid of this disclosure are, if necessary, not bactericidal compounds.

[0150] Administration of the preferred enantiomers of the 2-hydroxycarboxylic acid of this disclosure may, as necessary, cause dysregulation (suppression or enhancement) of the metabolism of planktonic bacteria, thereby being sufficient to allow other mechanisms (such as host defense mechanisms or bactericidal compounds) to have an effect of killing the planktonic bacteria whose metabolism has been dysregulated. Dysregulation (suppression or enhancement) of bacterial metabolism in one direction when the preferred enantiomers of the 2-hydroxycarboxylic acid are provided alone may be followed by dysregulation (enhancement or suppression) in the other direction when the preferred enantiomers of the 2-hydroxycarboxylic acid are provided in combination with antimicrobial compounds.

[0151] Planktonic bacteria administered with a preferred enantiomer of the 2-hydroxycarboxylic acid of this disclosure may be resistant to one or more antimicrobial compounds. The resistance mechanism may take the form of one or more of the following activity classes: - Prevents antimicrobial compounds from entering bacterial cells; - Altering the structure of the target of antimicrobial compounds; - Increasing the outflow of antimicrobial compounds into bacterial cells; and / or - Increases the metabolism of antimicrobial compounds within bacterial cells.

[0152] Dysregulation of metabolic activity induced by preferred enantiomers of the 2-hydroxycarboxylic acid of this disclosure can aid in the sensitization of planktonic bacteria to antimicrobial compounds (e.g., in the form of antibiotics). This dysregulation may be a suppression of bacterial metabolism, which may result in, for example, prevention of the extracellular efflux of antimicrobial compounds and / or a decrease in the metabolism of antimicrobial compounds within bacterial cells. Alternatively, if the dysregulation is an enhancement of bacterial metabolism induced by preferred enantiomers of the 2-hydroxycarboxylic acid of this disclosure, this may aid in sensitizing planktonic bacteria to antimicrobial compounds by, for example, increasing the ability of antimicrobial compounds to penetrate planktonic bacteria, increasing the amount of intracellular free radicals to make cells sensitive to antimicrobial compounds, and / or increasing the ability of antimicrobial compounds to bind to their targets, even if planktonic bacteria alter the target structure of the antimicrobial compound.

[0153] Methods for metabolic dysregulation This disclosure further relates to a method for dysregulating the metabolism of planktonic bacteria, the method being i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to floating bacteria.

[0154] This disclosure further relates to a method for dysregulating the metabolism of planktonic bacteria, the method being i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, a preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0155] This invention provides a method for treating or preventing bacterial infections caused by airborne bacteria, and this method is i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to the infection site.

[0156] This disclosure provides a method for treating or preventing bacterial infections caused by airborne bacteria, and this method is i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to the infection site, Here, a preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0157] This disclosure further relates to a method for sensitizing planktonic bacteria to an antimicrobial compound, the method being i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0158] This disclosure further relates to a method for sensitizing planktonic bacteria to an antimicrobial compound, the method being i. The process includes administering a preferred enantiomer of 2-hydroxycarboxylic acid to a floating bacterium. Here, the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0159] Preferably, dysregulation of the metabolism of planktonic bacteria is suppression of their metabolism. When a preferred enantiomer of 2-hydroxycarboxylic acid is administered in combination with an antimicrobial compound, the dysregulation may be suppression and subsequent enhancement of the metabolism of planktonic bacteria. However, in some cases, enhancement of the metabolism of planktonic bacteria may occur upon exposure to the preferred enantiomer of 2-hydroxycarboxylic acid alone, without being preceded by suppression and / or the presence of an antimicrobial compound. In such cases, metabolic suppression may follow the metabolic enhancement in the presence of both 2-hydroxycarboxylic acid and the antimicrobial compound.

[0160] The term "infection" includes the growth of microorganisms on and within surfaces, tissues, fluids, and gases, both biological and non-biological. For example, an infection could be the growth of Pseudomonas in lung tissue, an infection of Erwinia amylovora (bacterial burn disease) in apple and pear trees, the growth of Legionella in water pipes, or a bacterial infection of the blood (sepsis).

[0161] The compositions of this disclosure may contain 0.001% to 100% of a preferred enantiomer of 2-hydroxycarboxylic acid. For example, an ophthalmic composition may contain 0.001% to 1% of a preferred enantiomer of 2-hydroxycarboxylic acid. Washing and immersion, bandage and wipe, topical, inhalation, or oral compositions may contain 0.2% to 100% of a preferred enantiomer of 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient, and / or other activators. Optionally, a preferred enantiomer of 2-hydroxycarboxylic acid is D-lactic acid.

[0162] The compositions of this disclosure may contain undesirable enantiomers of 2-hydroxycarboxylic acid, the percentage of which may be less than 20%. Alternatively, the compositions of this disclosure may not contain undesirable enantiomers of 2-hydroxycarboxylic acid. Therefore, the compositions may contain only preferred enantiomers of 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0163] This disclosure further provides compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid for treating or preventing bacterial infections in a subject, wherein the bacterial infection is caused by planktonic bacteria, and the composition dysregulates the metabolism of the planktonic bacteria.

[0164] The Disclosure further provides compositions for treating or preventing infections in a subject, comprising a preferred enantiomer of a 2-hydroxycarboxylic acid in combination with an antimicrobial compound, wherein the infection is caused by a planktonic bacterium, and the composition dysregulates the metabolism of the planktonic bacterium.

[0165] The Disclosure further provides compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid for treating or preventing bacterial infections in a subject, wherein the bacterial infections are caused by planktonic bacteria, and the compositions sensitize the planktonic bacteria to an antimicrobial compound, wherein the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacteria.

[0166] The Disclosure further provides compositions comprising preferred enantiomers of 2-hydroxycarboxylic acid for use in combination with an antimicrobial compound to treat or prevent an infection in a subject, wherein the infection is caused by a planktonic bacterium, the composition sensitizes the planktonic bacterium to the antimicrobial compound, and the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacterium.

[0167] The effectiveness of a method of treating or preventing infection in a subject by administering a preferred enantiomer of 2-hydroxycarboxylic acid can be established by determining outcomes other than measuring the metabolism of the bacteria themselves. For example, effectiveness may be determined in a subject with infectious lung disease by a reduction in the bacterial load in a sputum sample after administration of the preferred enantiomer of 2-hydroxycarboxylic acid. In another example, the effectiveness of treatment using a preferred enantiomer of 2-hydroxycarboxylic acid for infection in the mammary gland may be established by a reduction in visible inflammation on the outer surface of the mammary gland tissue, or a reduction in swelling, fever, pain, and / or redness. In either the lung or mammary gland example, further testing may involve a microbiological evaluation of the biological fluid from the site of infection (e.g., sputum, breast milk) to determine the microbial load. If the preferred enantiomer of 2-hydroxycarboxylic acid dysregulates the metabolism of the parasitic bacteria causing the infection, the microbial load at the site of infection should be reduced.

[0168] surface Compositions comprising preferred enantiomers of the 2-hydroxycarboxylic acids of this disclosure may be used to dysregulate the metabolism of planktonic bacteria on living or non-living surfaces, and in living or non-living materials such as liquids and gases, and, if necessary, to sensitize planktonic bacteria to antimicrobial compounds. For the purposes and to facilitate the discussion of the present invention, the presence of planktonic bacteria in liquids and gases is considered to be "on the surface".

[0169] Planktonic bacteria can be present on or within almost any type of surface, including organic surfaces (living or non-living), plastics, metals, glass, soil particles, wood, and food. They can also be present in living or non-living fluids and gases such as blood, air, lymph, and drinking water. This disclosure may be used to dysregulate the metabolism of planktonic bacteria in such locations.

[0170] Biological surfaces that can host (host) planktonic bacteria capable of causing metabolic dysregulation using the compositions of this disclosure include: lung mucosal surfaces (particularly the lungs of subjects with cystic fibrosis or bronchiectasis); chronic wounds (including ulcers such as diabetic ulcers); skin surfaces (particularly those with cutaneous dysbiosis, acne, itchy skin, warts, psoriasis and atopic dermatitis); vaginal interiors (including bacterial vaginosis and vulvovaginal candidiasis); gastrointestinal surfaces (e.g., intestinal bacterial overgrowth, Clostridium Difficile infections, gastric ulcers), middle ear surfaces (especially those with otitis media); periodontal surfaces (especially those with periodontitis); paranasal sinus surfaces (especially those infected with sinusitis); cardiac surfaces (especially those with infectious endocarditis); ocular surfaces (especially those with postoperative endophthalmitis, microbial keratitis, infectious crystalline keratopathy); other ocular surfaces (especially those associated with scleral buckle insertion, conjunctival plug insertion, and lacrimal duct intubation); and bone surfaces (especially those with osteomyelitis). Biomaterials that can harbor planktonic bacteria whose metabolism can be inhibited using the compositions of this disclosure include blood, mucus, gastric juice, tears, and lymph.

[0171] The presence of airborne bacteria is common in the food industry, and since these bacteria can be pathogenic, it is a cause for concern.

[0172] Non-biological surfaces in the medical field that can harbor floating bacteria whose metabolism can be inhibited using the compositions of the Disclosure include urinary catheters, intravenous catheters, intrauterine devices, prostheses (including artificial hip, knee, and heart valves), cochlear implants, intraocular lenses, breast implants, vascular access devices, endotracheal tubes, tracheostomies, enteral feeding tubes, wound drains, ear canals, soap dispensers, and surface medical devices such as contact lenses, orthodontic retainers, and mouthguards. Non-biological materials in the medical field that can harbor floating bacteria whose metabolism can be dysregulated using the compositions of the Disclosure include oxygen or other air sources to assist the respiration of subjects, as well as handwashing solutions.

[0173] Industrial surfaces that can harbor planktonic bacteria capable of metabolic dysregulation include hydroelectric turbines, oil and gas pipelines, air conditioning units, soap dispensers, industrial water systems, ship hulls, and dairy farming equipment. Planktonic bacteria may also be present on animal habitats or materials and equipment related to livestock farming. Industrial substances that can harbor planktonic bacteria capable of metabolic dysregulation using the compositions of this disclosure include lubricating oils, oxygen or other air sources, and handwashing solutions.

[0174] plant Preferred enantiomers of the 2-hydroxycarboxylic acid compositions of this disclosure can be used to dysregulate the metabolism of planktonic bacteria in and on plant tissues, as well as in the soil in which plants grow, and, if necessary, to sensitize the planktonic bacteria to antimicrobial compounds. Many plant infections are treated with antimicrobial compounds, and resistance to antimicrobial compounds arises in a manner similar to that encountered in infections in animals. Resistance can be counteracted by sensitizing planktonic bacteria to antimicrobial compounds using preferred enantiomers of the 2-hydroxycarboxylic acid of the present invention.

[0175] Plants can include large-scale cultivated crops, vegetable crops, fruit crops, plantations, orchards, potted plants, ornamental gardens, parks, forests, and lawns.

[0176] Preferred enantiomers of the 2-hydroxycarboxylic acid compositions of this disclosure can be used to dysregulate the metabolism of planktonic bacteria on food crops in fields, orchards, vineyards and nurseries, large-scale cultivated crops, and vegetable and fruit crops, as well as on plants in home garden environments, potted plants, lawns, and sports fields. Furthermore, the 2-hydroxycarboxylic acid compositions can be used to dysregulate the metabolism of planktonic bacteria in intensive plant cultivation environments such as greenhouses. The 2-hydroxycarboxylic acid compositions can also be used to dysregulate the metabolism of planktonic bacteria in environments with few plants but high levels of bacteria, such as backyards, around pig farms, chicken coops and poultry houses, and barns.

[0177] Plant bacterial infections that can be treated with preferred enantiomers of the 2-hydroxycarboxylic acid compositions of this disclosure include infections caused by bacteria such as those of the genera Agrobacterium, Xanthomonas, Pseudomonas, Erwinia, Corynebacterium, Ralstonia, and Streptomyces. For example, the following diseases can be treated: [Table 7]

[0178] delivery The compositions of this disclosure may be provided, for example, as topical compositions such as ointments, creams, foams, adhesives or lotions, eye ointments and eye drops or ear drops, or nasal sprays; as internal topical compositions such as impregnated bandages, patches and wipes; as pessaries and suppositories; as inhalation compositions such as sprays or dry powder compositions; as injectable formulations for intravenous and local injection (e.g., into abscesses); or as oral formulations such as tablets, liquid capsules, or oral gels. The compositions may further be in the form of washing solutions or immersion solutions. The compositions may further be in the form of foliar sprays or soil sprays.

[0179] As used herein, the term “therapeutic dose” means an amount of the composition sufficient to at least partially achieve the desired effect when administered according to the desired dosing regimen or application regimen. Where necessary, the desired effect is dysregulation of the metabolism of planktonic bacteria. However, the effect may be to delay, inhibit, or partially or completely halt the onset or progression of infections caused by planktonic bacteria (including infections of animals, surfaces, or plants).

[0180] As used herein, the term “prophylactic effective dose” means an amount of a composition sufficient to cause at least partially dysregulated metabolism of a planktonic bacterium when administered according to the desired dosing regimen or application regimen.

[0181] Metabolic dysregulation in planktonic bacteria in the presence of the preferred enantiomer of 2-hydroxycarboxylic acid alone may result in either suppression or enhancement of the planktonic bacteria's metabolism. Following the introduction of an antimicrobial compound, the dysregulation (suppression or enhancement) may persist, possibly with an increased degree of dysregulation. However, when planktonic bacteria are present with both the preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, the metabolic dysregulation may change from previously suppressive to enhanced, and from previously enhanced to suppressive. Alternatively, metabolic dysregulation may not occur in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation may occur in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, particularly if the planktonic bacteria were resistant to antimicrobial compounds in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid.

[0182] Based on the above, it will be understood by those skilled in the art that a single subject or surface can be treated using multiple different treatment and administration methods. For example, a subject already receiving pharmaceuticals (e.g., intravenous antibiotics) may benefit from oral delivery, intravenous delivery, inhalation, or topical application of the compositions of this disclosure. Some subjects may be administered only the compositions containing a preferred enantiomer of 2-hydroxycarboxylic acid by oral administration, inhalation, or topical application. For example, a subject may have symptoms of cystic fibrosis, be diagnosed with a pulmonary infection, or have symptoms of a medical condition, and these symptoms may benefit from administration to the subject of an inhalation composition containing a preferred 2-hydroxycarboxylic acid. Alternatively, a subject may have a local infection such as a chronic wound, an eye infection, or periodontitis, and may be administered topically with a preferred enantiomer of 2-hydroxycarboxylic acid. A subject may have intestinal disruption ("dysbiosis") (e.g., small intestine) which may benefit from oral administration of a preferred enantiomer of the 2-hydroxycarboxylic acid composition of this disclosure. These dysbiosis may contribute to enteropathogenic disorders, inflammatory bowel disease (IBS), Crohn's disease, ulcerative colitis, or colorectal cancer.

[0183] The compositions of this disclosure may be used in plants and in the soil in which the plants grow to treat plant infections on and within plant tissues.

[0184] The compositions of this disclosure may be used as cleaning or immersion solutions for non-living surfaces such as indwelling equipment and water pipes, in the form of wet wipes, on non-living surfaces, and as cleaning compositions for pipes, food processing equipment and medical devices. When the compositions are applied to a non-living surface, cleaning, washing, or immersion with 2-hydroxycarboxylic acid may be followed by cleaning with conventional detergents, UV sterilization, or bleaching.

[0185] The compositions of this disclosure may also be used diagnostically. In one embodiment, for example, a subject may receive a dose of the compositions of this disclosure as part of a procedure for diagnosing an infection associated with airborne bacteria (such as a pulmonary infection), in which one of several symptoms of the subject is improved according to the composition.

[0186] According to a particular embodiment, the composition is administered periodically until the treatment is achieved. In one preferred embodiment, the composition is administered to a subject requiring such treatment using a dosing regimen selected from the group consisting of every hour, every two hours, every three hours, once daily, twice daily, three times daily, four times daily, five times daily, once weekly, twice weekly, once every two weeks, and once monthly. However, other application schedules may be used in accordance with this disclosure. If necessary, the composition of the treatment regimen is administered to the subject one to five times a day, for example, once or twice a day.

[0187] When administration is to a non-living surface, the composition may be applied to the surface periodically until the removal or destruction of planktonic bacteria is achieved, as necessary, by dysregulation of the planktonic bacteria' metabolism. In one preferred embodiment, the composition is applied to the surface every hour, every two hours, every three hours, once daily, twice daily, three times daily, four times daily, five times daily, once weekly, twice weekly, once every two weeks, and once monthly. However, other application schedules may be utilized in accordance with this disclosure. The composition can be applied to the implanted device before insertion and after removal.

[0188] If necessary, the compositions of this disclosure may be administered, for example, orally, topically (ocular, buccal, and sublingual, rectally, vaginally, intranasally), or as an aerosol. When the compositions are delivered to a non-living surface, they may be administered as a wash, immersion solution, or wipe. The mode of administration or dispensing may, if necessary, be appropriate to the form in which the composition is prepared. The mode of administration for the most effective response may be determined empirically, and the means of administration or dispensing described below are given as examples and are not in any way limiting the methods of delivery of the compositions of this disclosure.

[0189] The compositions of this disclosure may optionally include pharmaceutically acceptable non-toxic excipients and carriers. As used herein, “pharmaceutically acceptable carrier” is a pharmaceutically acceptable solvent, suspending agent, excipient, or vehicle for delivering the compound to a target. The carrier may be liquid or solid and is selected with the planned mode of administration in mind.

[0190] The compositions of this disclosure may be selected from the group consisting of immediate-release compositions, delayed-release compositions, controlled-release compositions, and rapid-release compositions.

[0191] The compositions of this disclosure may further include an anti-inflammatory agent (such as a corticosteroid).

[0192] The compositions described herein may be formulated as oil-in-water emulsions and / or water-in-oil emulsions. In such compositions, the immediate-release dosage form may be in a continuous phase, and the delayed-release dosage form may be in a discontinuous phase. The compositions may also be manufactured in a manner for the delivery of the three dosage forms as described above herein. For example, an oil-in-water emulsion may be provided having an oil that is a continuous phase containing an immediate-release component, water dispersed in the oil containing a first delayed-release dosage form, and oil dispersed in the water containing a third delayed-release dosage form.

[0193] The compositions described herein may be in the form of liquid compositions. Liquid compositions may include solutions containing a therapeutic agent dissolved in a solvent. Generally, any solvent can be used in which the therapeutic agent dissolves and which has the desired effect that can be administered to a target. Generally, any concentration of the therapeutic agent that has the desired effect can be used. In some variations, the compositions are solutions that are unsaturated, saturated, or supersaturated. The solvent may be a pure solvent or a mixture of liquid solvent components. In some variations, the formed solution is an in-situ gelled composition. The types of solvents and solutions that can be used are well known to those familiar with such drug delivery techniques.

[0194] This composition may or may not contain water. If necessary, the composition contains water, i.e., is aqueous. In another embodiment, this composition does not contain preservatives. In one embodiment, the composition described herein may be aqueous and may contain 0 to 90% water. In another embodiment, the aqueous composition described herein may contain 20 to 80% water. In yet another embodiment, the aqueous composition may contain 50 to 70% water. The water may further include ordinary water, distilled water, sterile water, demineralized water, or deionized water. Alternatively, the composition may be non-aqueous and contain no water or a negligible amount of water (e.g., less than 1%, less than 0.1%, less than 0.01%).

[0195] Pharmaceutical or veterinary compositions may be formulated according to conventional pharmaceutical or veterinary practices (see, for example, Remington: The Science and Practice of Pharmacy, 20th edition, 2000, ed; ARGennaro, Lippincott Williams & Wilkins, Philadelphia, and Encyclopedia of Pharmaceutical Technology, eds; J. Swarbrick and JCBoylan, 1988–1999, Marcel Dekker, New York; Remington's Pharmaceutical Sciences, 18th edition, Mack Publishing Company, Easton, Pennsylvania, USA).

[0196] Generally, suitable carriers, excipients, and diluents include, but are not limited to, water, physiological saline, ethanol, dextrose, glycerol, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water syrup, methylcellulose, methyl and propyl hydroxybenzoate, polysorbate, talc, magnesium stearate, mineral oil, or combinations thereof. The composition may further contain lubricants, pH buffers, wetting agents, emulsifiers and suspending agents, preservatives, sweeteners, or flavoring agents.

[0197] The composition may be in the form of a controlled-release composition and may include a degradable or non-degradable polymer, a hydrogel, an organogel, or other physical constructs that modify the release of a preferred enantiomer of a 2-hydroxycarboxylic acid. It is understood that such a composition may include additional inert components added to provide a desired color, stability, buffering capacity, dispersion, or other known desirable characteristics. Such a composition may further include liposomes such as emulsions, foams, micelles, insoluble monolayers, liquid crystals, phospholipid dispersions, and lamellar layers. Liposomes for use in this disclosure may generally be formed from standard vesicle-forming lipids, including neutral and negatively charged phospholipids and sterols, such as cholesterol.

[0198] Washing and immersion compositions Compositions of the present disclosure comprising a preferred enantiomer of 2-hydroxycarboxylic acid can be used to clean and / or immerse equipment such as indwelling medical devices or water piping systems. Cleaning or immersing such devices with a composition comprising a preferred enantiomer of 2-hydroxycarboxylic acid assists in sterilizing or cleaning the devices by helping to dysregulate the metabolism of any floating bacteria present on the device surface, and thus making it easier to remove and / or kill any microorganisms present. The treatment may, if necessary, dysregulate metabolism sufficiently to increase the effectiveness of antimicrobial compounds (e.g., bactericidal, bacteriostatic, antibiofilm, or disinfectant antimicrobial compounds).

[0199] In one embodiment, the composition used for washing or immersion may contain 0.1% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient, and / or other activators.

[0200] In one embodiment, the composition used for washing or immersion may contain an undesirable enantiomer of 2-hydroxycarboxylic acid, the percentage of which may be less than 20%. Alternatively, the composition of the present disclosure may not contain an undesirable enantiomer of 2-hydroxycarboxylic acid. Therefore, the composition may contain only a preferred enantiomer of 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0201] Bandages and wipes The compositions of the present disclosure, comprising preferred enantiomers of 2-hydroxycarboxylic acid, may be provided in the form of impregnated dressings or bandages, or moistened wipes. Dressings may be in a form that can be applied to wounds or local surfaces that may be infected with planktonic bacteria, and may be left in situ for several hours, several days, or several weeks. Wipes can be used to reduce the presence of planktonic bacteria, for example, on wounds or mucosal surfaces, such as vaginal or rectal surfaces, or on non-living surfaces. Wipes may be used hourly, daily, weekly, or as needed. For example, wipes may be used after each urination or defecation to dysregulate the metabolism of planktonic bacteria present on vaginal or rectal mucosal tissue. Alternatively, wipes may be used on food preparation surfaces before use.

[0202] In one embodiment, the composition used for bandages or wipes may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may consist of a carrier, diluent or excipient, and / or other activators.

[0203] In one embodiment, a composition used for bandages or wipes may contain an undesirable enantiomer of 2-hydroxycarboxylic acid, the percentage of which may be less than 20%. Alternatively, the composition of the present disclosure may not contain an undesirable enantiomer of 2-hydroxycarboxylic acid. Therefore, the composition may contain only a preferred enantiomer of 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0204] inhalation delivery The compositions of this disclosure may be delivered by spray delivery or by other delivery devices. This is particularly suitable for airborne bacteria-related respiratory and ear-nose-throat (ENT) diseases.

[0205] If necessary, the composition is administered to the target subject approximately once to six times per day, for example, once or twice per day.

[0206] Alternatively, the composition may be administered to the target subject by continuous inhalation via a nebulizer. The spray composition may be delivered at 24-hour, 12-hour, 8-hour, 6-hour, 4-hour, 2-hour, or 1-hour intervals, and each of these deliveries (except for the 24-hour and 12-hour intervals) may be repeated several times within a 24-hour period.

[0207] In one embodiment, the composition delivered by nebulizer or aerosol delivery may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient, and / or other activators.

[0208] In one embodiment, the composition delivered by nebulizer or aerosol delivery may contain an undesirable enantiomer of 2-hydroxycarboxylic acid, the percentage of which may be less than 20%. Alternatively, the composition of the present disclosure may not contain an undesirable enantiomer of 2-hydroxycarboxylic acid. Therefore, the composition may contain only a preferred enantiomer of 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0209] Subjects may typically be administered ±20% or ±10% of a preferred enantiomer of 2-hydroxycarboxylic acid in doses of 1 to 500 mg / mL. Doses may be delivered over various periods, for example, in a single puff (seconds) or over several minutes to several hours. Doses may be delivered over 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 45 minutes, 60 minutes, 120 minutes, or longer. The dose is typically administered by a nebulizer or by at least one, preferably several “puffs,” from an aerosol device. For example, subjects may receive a preferred enantiomer of 2-hydroxycarboxylic acid between 1 mg / mL and 500 mg / mL in single doses per day or in several doses per day.

[0210] The total daily dose is administered at least once per day, if necessary, but may be divided into two or more doses per day. Some subjects may benefit from a “loading” period with preferred enantiomers of 2-hydroxycarboxylic acid, involving higher or more frequent doses over several days or weeks, followed by reduced or maintenance doses. Since cystic fibrosis, COPD, etc., are typically chronic conditions, subjects are expected to receive such therapy over a long period.

[0211] A wide range of mechanical devices designed for the pulmonary delivery of therapeutic products exist, including, but not limited to, nebulizers, metered-dose inhalers, and powder inhalers, all of which are well known to those skilled in the art. Some specific examples of commercially available devices suitable for carrying out this disclosure are the Ultravent nebulizer from Mallinckrodt, Inc., St. Louis, Missouri; the Acorn II nebulizer from Marquest Medical Products, Englewood, Colorado; the Ventolin metered-dose inhaler from Glaxo Inc., Research Triangle Park, North Carolina; and the Spinhaler powder inhaler from Fisons Corp., Bedford, Massachusetts.

[0212] All such devices require the use of compositions suitable for the distribution of preferred enantiomers of 2-hydroxycarboxylic acids. Typically, each composition is specific to the type of device used and may include the use of appropriate propellant materials in addition to common diluents, adjuvants, and / or carriers useful for the therapy.

[0213] Regardless of the form of the drug composition, it is preferable to produce droplets or particles for inhalation in the range of approximately 0.1 μm to 12 μm, or approximately 0.25 μm to 6 μm, and optionally 1 μm to 6 μm, for example, approximately 2 μm to 4 μm. Alternatively, the particles may be 0.1 μm to 1.0 μm, 0.2 μm to 0.9 μm, 0.3 μm to 0.8 μm, 0.4 μm to 0.7 μm, or 0.5 μm. By creating inhalation particles with a relatively narrow size range, it is possible to further increase the efficiency of the drug delivery system and improve the reproducibility of administration. Therefore, it is preferable that the particles not only have a size range of 0.1 μm to 12 μm or 2 μm to 6 μm or approximately 3 to 4 μm, but also that the average particle size is within a narrow range such that 80% or more of the particles delivered to the target have a particle size within ±20% of the average particle size, or within ±10% of the average particle size, for example, within ±5% of the average particle size.

[0214] "Particle size" is a concept introduced to compare the dimensions of solid particles and liquid particles (droplets). For droplets and aerosols, terms such as "aerodynamic diameter" and "mass-center aerodynamic diameter" (MMAD) are used.

[0215] The "aerodynamic diameter" is the diameter of a unit-density sphere that has the same terminal settling velocity as the particle in question. It is used to predict where such a particle will settle in the airway.

[0216] The "mass-center aerodynamic diameter" is the geometric mean aerodynamic diameter. 50% of particles (by weight) will have a MMAD smaller than the MMAD, and 50% will have a MMAD larger than the MMAD.

[0217] During particle sizing experiments, the suspension contains countless particles of various sizes in motion. When a particle sizing machine analyzes these particles, a particle distribution curve is formed, covering the entire particle size range from the smallest particles (which can be 1 nm) to the largest particles (which can be 100 μm). The cumulative frequency of each particle is calculated from the particle size distribution curve. 10 This refers to a specific particle diameter in which 10% of the particles in a suspension have a diameter smaller than or equal to a specific particle diameter.

[0218] D 50 :D 10 Similarly, D 50 This refers to the 50% cutoff diameter of the particle population in the composition, where 50% of the particles in the suspension have a diameter smaller than or equal to a specific particle diameter.

[0219] D 90 :D 90 This refers to the 90% cutoff diameter of the particle population in the composition, where 90% of the particles in the suspension have a diameter smaller than or equal to a specific particle diameter.

[0220] The term “airway” should be interpreted as meaning the system of cells and organs that function in respiration, and in particular the organs, tissues and cells of the airway include the lungs, nose, nasal tract, sinuses, nasopharynx, larynx, trachea, bronchi, bronchioles, respiratory bronchioles, alveolar ducts, alveolar sacs, alveoli, lung cells (type 1 and type 2), ciliated mucosal epithelium, mucosal epithelium, squamous epithelial cells, mast cells, goblet cells, neutrophils, macrophages and intraepithelial dendritic cells.

[0221] In one embodiment of the present disclosure, a method for dysregulating the metabolism of airborne bacteria in the target lung comprises administering a therapeutically effective concentration of an inhalation composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid in the form of a dose of 1 to 1000 mg / mL, for example, 5 to 500 mg / mL or greater.

[0222] In one embodiment of the present disclosure, a method for dysregulating the metabolism of airborne bacteria in the target lung comprises administering a therapeutically effective concentration of an inhalation composition comprising a preferred enantiomer of 2-hydroxycarboxylic acid in the form of one or more doses of 1 to 1000 mg / kg / day, for example, 6 to 600 mg / kg / day.

[0223] One embodiment of the present disclosure provides a method for dysregulating the metabolism of airborne bacteria in the lungs of a target by administering an inhalation composition containing a preferred enantiomer of 2-hydroxycarboxylic acid at a prophylactic effective concentration in the form of 1 to 1000 mg / mL, for example, 5 to 500 mg / mL or more.

[0224] The compositions of this disclosure may be administered to subjects using disposable packaging and portable, handheld, battery-powered devices such as AERx devices (U.S. Patent No. 5,823,178, Aradigm, Hayward, Calif.). Alternatively, the compositions of this disclosure may be administered using mechanical (non-electronic) devices. Other inhalation devices, including conventional jet nebulizers, ultrasonic nebulizers, soft mist inhalers, dry powder inhalers (DPIs), medium-dose inhalers (MDIs), condensed aerosol generators, and other systems, may be used to deliver the compositions.

[0225] For use as an aerosol, the compounds of the present disclosure in solution or suspension may be packaged in a pressurized aerosol container with a suitable propellant, such as a hydrocarbon propellant including a conventional adjuvant, such as propane, butane, or isobutane. A dry powder inhaler is a system that can operate with a pressurized air source to produce dry powder particles of a pharmaceutical composition compressed to a very small volume. For inhalation, the system has a plurality of chambers or blisters, each containing a single dose of the pharmaceutical composition and a select element for releasing a single dose.

[0226] Aerosols can be generated by forcing a drug through pores in a membrane having a size in the range of about 0.25–6 μm (U.S. Patent No. 5,823,178). If the pores are of this size, the particles escaping through the pores to generate the aerosol will have a diameter in the range of 0.5–12 μm. Drug particles can be released with an airflow intended to keep the particles within this size range. The generation of small particles can be facilitated by the use of a vibrating device providing an vibration frequency in the range of about 800–4000 kilohertz. Those skilled in the art will recognize that, keeping in mind that the objective of some embodiments is to provide aerosolized particles having a diameter in the range of about 0.5–12 μm, several adjustments can be made to parameters such as the size of the pores through which the drug is released, the vibration frequency, the pressure, and other parameters based on the density and viscosity of the composition.

[0227] Local delivery The compositions of this disclosure may be delivered topically. Topical administration may involve directly administering a therapeutically effective amount of a preferred enantiomer of 2-hydroxycarboxylic acid to the skin, eye, or mucous membrane surface of the subject. If necessary, the preferred enantiomer of 2-hydroxycarboxylic acid is applied topically to the skin, mucous membrane (oral, nasal, vaginal, rectal) or eye of the subject. Use may also involve administering a therapeutically effective amount of a preferred enantiomer of 2-hydroxycarboxylic acid to the skin, mucous membrane (oral, nasal, vaginal, rectal) or eye of the subject.

[0228] The compositions of this disclosure can be administered topically. Therefore, the use described herein is intended for compositions suitable for direct application to the skin.

[0229] In one embodiment, the composition used for topical delivery may contain 0.2% to 100% of a preferred enantiomer of a 2-hydroxycarboxylic acid. The remainder of the composition may consist of a carrier, diluent or excipient, and / or other activators.

[0230] In one embodiment, the composition used for local delivery may contain an undesirable enantiomer of 2-hydroxycarboxylic acid, the percentage of the undesirable enantiomer of 2-hydroxycarboxylic acid may be less than 20% of the total 2-hydroxycarboxylic acid content. Alternatively, the composition of the present disclosure may not contain an undesirable enantiomer of 2-hydroxycarboxylic acid. Therefore, the composition may contain only a preferred enantiomer of 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0231] The composition may be in a form selected from the group including suspensions, emulsions, liquids, creams, oils, lotions, ointments, gels, hydrogels, pastes, plasters, roll-on liquids, skin patches, sprays, glass bead bandages, synthetic polymer bandages, and solids. For example, the compositions of the present disclosure may be provided in the form of aqueous compositions or ointments based on organic solvents such as oils. Alternatively, the compositions of the present disclosure may be provided by a liquid spray containing at least a solvent in which a film-forming component and a preferred enantiomer of 2-hydroxycarboxylic acid are dispersed or solubilized.

[0232] The compositions of this disclosure may be provided in forms selected from the group including, but are not limited to, rinses, shampoos, lotions, gels, leave-on formulations, wash-off formulations, and ointments.

[0233] Depending on the preferred treatment regimen, various topical delivery systems may be suitable for administering the compositions of the present disclosure. Topical compositions can be prepared by dissolving or combining preferred enantiomers of the 2-hydroxycarboxylic acids of the present disclosure in an aqueous or non-aqueous carrier. Generally, any liquid, cream, gel, or similar substance that does not react to the compound or any other active ingredient to a recognizable degree and is non-irritating is suitable and can be introduced into the composition. Suitable non-sprayable, semi-solid, or solid forms containing a carrier suitable for topical application and having a kinematic viscosity greater than water may also be used.

[0234] Suitable compositions are well known to those skilled in the art and include, but are not limited to, solutions, suspensions, emulsions, creams, gels, ointments, powders, liniments, plasters, aerosols, and transdermal patches, which may be sterilized as desired or mixed with auxiliary agents such as preservatives, stabilizers, emulsifiers, humectants, fragrances, colorants, odor control agents, thickeners, such as natural rubber. Particularly preferred topical compositions include ointments, creams, or gels.

[0235] Ointments are generally prepared using either (1) an oily base, i.e., a fixed oil or hydrocarbon, such as white petroleum or mineral oil, or (2) an absorbent base, i.e., an anhydrous substance or a substance capable of absorbing water, such as anhydrous lanolin. Typically, after the base is formed, a preferred 2-hydroxycarboxylic acid is added to give the desired concentration, whether oily or absorbent.

[0236] Creams are oil / water emulsions. They typically consist of an oil phase (internal phase) containing fixed oils, hydrocarbons, waxes, petroleum, mineral oil, etc., and an aqueous phase (continuous phase) containing water and any water-soluble substances, such as added salts. The two phases are stabilized by the use of emulsifiers, such as surfactants like sodium lauryl sulfite; or hydrophilic colloids such as acacia colloidal clay or vegetable gums like bee gum. During emulsion formation, a preferred enantiomer of 2-hydroxycarboxylic acid can be added in an amount that achieves the desired concentration.

[0237] The gel comprises a base selected from an oily base, water, or an emulsion-suspension base. A gelling agent is added to the base to form a matrix within the base and increase its viscosity. Examples of gelling agents include hydroxypropyl cellulose and acrylic acid polymers. Typically, a preferred enantiomer of 2-hydroxycarboxylic acid is added to the composition at a desired concentration prior to the addition of the gelling agent.

[0238] Topically delivered compositions for application to mucosal surfaces (e.g., oral mucosal surfaces, vaginal mucosal surfaces, nasal mucosal surfaces, or rectal mucosal surfaces, etc.) or skin wounds may contain 5% to 100% of a preferred enantiomer of 2-hydroxycarboxylic acid. The remainder of the composition may consist of a carrier, diluent or excipient, and / or other activators. In one embodiment, higher doses may be used in medically controlled situations, and lower doses may be used for non-life-threatening wounds treated at home.

[0239] In one embodiment, a composition used for application to a mucosal surface and / or skin wound may contain an undesirable enantiomer of 2-hydroxycarboxylic acid, the percentage of which may be less than 20%. Alternatively, the composition of this disclosure may not contain an undesirable enantiomer of 2-hydroxycarboxylic acid. Therefore, the composition may contain only a preferred enantiomer of 2-hydroxycarboxylic acid as the 2-hydroxycarboxylic acid.

[0240] The compositions of the present disclosure can be administered by ocular topical delivery. If necessary, the ophthalmic composition contains a preferred enantiomer of 2-hydroxycarboxylic acid in an amount of 0.001% to 1%.

[0241] Ocular delivery includes delivery to the sclera, retina, intraocular fluid, and tissues around the eye. For example, the delivery can be topical delivery (cream, gel, ointment, spray, eye drops), an intraocular implant or other means.

[0242] Ocular delivery may also include injecting a preferred enantiomer of 2-hydroxycarboxylic acid into the sclera, the eye cavity or the area behind the eye. Compositions suitable for ocular injection include, if necessary, a sterile aqueous solution (in the case of water solubility) or dispersion, and a sterile powder for the immediate preparation of a sterile injection solution or dispersion. Alternatively, in certain embodiments, the compounds of the present disclosure are encapsulated in liposomes and delivered in an injectable solution to assist their transport across cell membranes. In an alternative or in addition, such preparations contain components of self-assembling pore structures to facilitate transport across cell membranes.

[0243] Oral delivery composition The compositions of the present disclosure can be administered by oral delivery.

[0244] In one form, the composition for oral delivery may contain 5% to 100% of a preferred enantiomer of 2-hydroxycarboxylic acid. The remainder of the composition may contain a carrier, diluent or excipient, and / or other active agents.

[0245] In one form, the composition for oral delivery may contain a less preferred enantiomer of 2-hydroxycarboxylic acid, and the % of the less preferred enantiomer of 2-hydroxycarboxylic acid may be less than 20%. Alternatively, the compositions of the present disclosure may not contain a less preferred enantiomer of 2-hydroxycarboxylic acid. Thus, the composition may contain only the preferred enantiomer of 2-hydroxycarboxylic acid as 2-hydroxycarboxylic acid.

[0246] The subjects may typically be administered a preferred enantiomer (+20% or +10%) of 2-hydroxycarboxylic acid at doses of 0.1 mg / kg / day to 2 g / kg / day.

[0247] Despite the varying pH levels of the digestive system (acidic in the stomach, alkaline in the small intestine), the efficacy of the preferred enantiomer of 2-hydroxycarboxylic acid against planktonic bacteria remains. The pH of the surrounding fluid does not affect the metabolic dysregulation ability of the preferred enantiomer of 2-hydroxycarboxylic acid. This is because 2-hydroxycarboxylic acid has been shown to remain stable in the gastrointestinal tract and maintain its functionality, which has been observed in numerous studies investigating acid resistance in lactic acid bacteria (e.g., Wang et al., 2018, Archives of Microbiology, 200, 195-201) and lactic acidosis (metabolic acidosis resulting from excessive fermentation in the gastrointestinal tract).

[0248] The oral compositions of this disclosure may be delivered together with immunostimulants or modifiers. Examples of immunostimulants or modifiers include specific immunostimulants (such as vaccines and antigens) and nonspecific agents, such as bee products (including propolis and honey), probiotics and prebiotics, hormones, vitamins (vitamin C, vitamin D), minerals (zinc oxide), antioxidants (including glutathione), interferons (including INF-alpha), interleukins (including interleukin-10), colchicine, thalidomide, and imiquimod. The immune booster may be delivered simultaneously with or after the composition containing the preferred enantiomer of the 2-hydroxycarboxylic acid.

[0249] plant delivery The composition containing a preferred enantiomer of 2-hydroxycarboxylic acid can be applied to plants and / or soil in the form of spraying using a pump pack, hand sprayer or boom sprayer, or in the form of application using a hose, jet, or aircraft equipped with a pesticide sprayer, and can also be applied through irrigation systems, i.e., drip, small sprinklers, flood irrigation, etc. The composition can also be further injected into plants or trees.

[0250] subject When a preferred enantiomer of 2-hydroxycarboxylic acid is administered to a subject, the subject may be any subject capable of infection by bacteria, viruses, fungi, or archaea. The subject may be any animal, including fish, crustaceans, mollusks, mammals, reptiles, or birds. If necessary, the subject may be a mammal, selected from the group including humans, dogs, birds, pigs, cattle, sheep, horses, rodents, weasels, rabbits, and cats. The subject may be a companion animal, livestock, or an animal of aquaculture or agricultural importance. In one example, the subject may be a human.

[0251] Excipients The forms of the compositions described herein that are illustrated above can be manufactured by methods well known to those skilled in the art of compositional science. Furthermore, the compositions described herein may optionally contain other excipients to assist in the manufacture and / or administration of the compositions described herein. In one embodiment, the composition further comprises one or more pharmaceutically or physiologically acceptable carriers, diluents or excipients. Non-limiting examples of such excipients, which are well known in the art, include flavorings, colorants, palates, antioxidants, viscosity modifiers, isotonic agents, drug carriers, sustained-release agents, comfort enhancers, emulsifiers, solubilizers, lubricants, binders and other stabilizers to assist in the manufacture and / or administration of the compositions.

[0252] The compositions disclosed herein are intended for use in pharmaceutical or veterinary applications, or in cleaning equipment such as medical devices, water pipes, or food preparation equipment.

[0253] If necessary, the composition is sterile. In addition to, or instead of, sterilization, the compositions of this disclosure may contain pharmaceutically acceptable preservatives to minimize the possibility of microbial contamination. In another embodiment, the compositions of this disclosure are stable. Pharmaceutically acceptable preservatives may be used in the composition to enhance the stability of the composition. However, it should be noted that, since the treated tissue may be sensitive to irritants, any preservative must be selected for safety. Suitable preservatives for use herein include, but are not limited to, phenylethyl alcohol, benzalkonium chloride or benzoic acid, or benzoates such as sodium benzoate and phenylethyl alcohol, which protect the solution from contamination by pathogens. In certain embodiments, the compositions of this disclosure contain about 0.001% to about 10.0% w / w benzalkonium chloride, or about 0.01% v / w phenylethyl alcohol. The preservative may also be present in an amount of approximately 0.001% to 1%, for example, approximately 0.002% to 0.02%, or approximately 0.02% w / w.

[0254] The compositions provided herein may also contain one or more emulsifiers, wetting agents, or suspending agents in amounts of about 0.001% to about 90%, or about 0.001% to about 50%, or about 0.001% to about 25%, or about 0.001% to about 10%, or about 0.001% to about 1%.Such agents as used herein include, but are not limited to, polyoxyethylene sorbitan fatty acids or polysorbates, including polyethylene sorbitan monooleate (polysorbate 80), polysorbate 20 (polyoxyethylene (20) sorbitan monolaurate), polysorbate 65 (polyoxyethylene (20) sorbitan tristearate), polyoxyethylene (20) sorbitan monooleate, polyoxyethylene (20) sorbitan monopalmitate, and polyoxyethylene (20) sorbitan monostearate; lecithin; agar; carrageenan; locust bean gum; guar gum; tragacanth; acacia; xanthan gum; karaya gum; pectin; amidated pectin; ammonium phosphatide; microcrystalline cellulose; methylcellulose; hydroxypropylcellulose; hydroxypropyl methylcellulose; ethyl methylcellulose; carboxymethylcellulose; sodium, potassium, and calcium salts of fatty acids; mono - and diglycerides; acetate esters of mono- and diglycerides of fatty acids; lactate esters of mono- and diglycerides of fatty acids; citrate esters of mono- and diglycerides of fatty acids; tartrate esters of mono- and diglycerides of fatty acids; mono- and diacetyltartrate esters of mono- and diglycerides of fatty acids; mixed acetate and tartrate esters of mono- and diglycerides of fatty acids; sucrose esters of fatty acids; sucrose esters; polyglycerol esters of fatty acids Examples include sterol; polyglycerol esters of polycondensed fatty acids of castor oil; propane-1,2-diol esters of fatty acids; sodium stearoyl-21-actylate; calcium stearoyl-2-lactate; stearoyl tartrate; sorbitan monostearate; sorbitan tristearate; sorbitan monolaurate; sorbitan monooleate; sorbitan monopalmitate; quillaja extract; polyglycerol esters of dimerized fatty acids of soybean oil; oxidatively polymerized soybean oil; and pectin extract.

[0255] The compositions of this disclosure may contain about 0.001% to about 5% (by weight) of a humectant to inhibit drying of mucous membranes and prevent irritation. For example, any of a variety of pharmaceutically acceptable humectants, including sorbitol, propylene glycol, polyethylene glycol, glycerol, or mixtures thereof, can be used.

[0256] This disclosure encompasses variations in the above composition, as the amounts of each compound may vary by ±5%, ±7.5%, ±10%, ±15%, ±17.5%, or ±20%.

[0257] This disclosure encompasses compositions in which the relative proportions of the active ingredient and / or each excipient differ independently from those specified above. In one embodiment of this disclosure, the relative proportions of the active ingredient and / or each excipient vary independently up to 50% from those specified above. In one embodiment of this disclosure, the relative proportions of the active ingredient and / or each excipient vary independently up to 40% from those specified above. In one embodiment of this disclosure, the relative proportions of the active ingredient and / or each excipient vary independently up to 30% from those specified above. In one embodiment of this disclosure, the relative proportions of the active ingredient and / or each excipient vary independently up to 20% from those specified above. In one embodiment of this disclosure, the relative proportions vary independently up to 10% from those specified above. In one embodiment of this disclosure, the relative proportions of the active ingredient and / or each excipient vary independently up to 5% from those specified above. In one embodiment of this disclosure, the relative proportions may vary independently from those specified above, up to a maximum of 10%. In one embodiment of this disclosure, the relative proportions of the active ingredient and / or each excipient may vary independently from those specified above, up to a maximum of 2%.

[0258] As will be understood by those skilled in the art, the combined percentage of excipients and activators may not exceed 100%, and the modifications described above are subject to this limitation. As will be understood by those skilled in the art, since the forms of the disclosure include components other than those specified, the combined percentage of excipients and activators may be less than 100%.

[0259] The above fluctuations represent fluctuations in the percentage of the relative proportion. For example, a 20% fluctuation in the relative proportion of an ingredient (excipient or surfactant) specified as 1% means that the relative proportion of that ingredient could be between 0.8% and 1.2%.

[0260] Inhalation delivery excipients Compositions suitable for use in nebulizers (either jet or ultrasonic) typically contain a preferred enantiomer of 2-hydroxycarboxylic acid suspended in water or a non-aqueous solvent. The composition may also contain buffers and simple sugars (e.g., for osmotic pressure stabilization and regulation). Nebulizer compositions may also contain surfactants to reduce or prevent surface-induced aggregation of the preferred enantiomer of 2-hydroxycarboxylic acid caused by atomization of the solution during aerosol formation. The use of liposomes, microcapsules or microspheres, inclusion complexes, or other types of carriers is also conceivable.

[0261] Compositions for use with metered-dose inhalation devices generally comprise a fine powder containing a preferred enantiomer of a 2-hydroxycarboxylic acid suspended in a propellant using a surfactant. The propellant may be any conventional material used for this purpose, such as hydrocarbons including chlorofluorocarbons, hydrochlorofluorocarbons, hydrofluorocarbons, or trichlorofluoromethane, dichlorodifluoromethane, dichlorotetrafluoroethanol, and 1,1,1,2-tetrafluoroethane, or combinations thereof. Suitable surfactants include sorbitan trioleate and soy lecithin. Oleic acid may also be useful as a surfactant.

[0262] The composition for dispensing from the powder inhalation device comprises finely divided dry powder containing a preferred enantiomer of 2-hydroxycarboxylic acid, and may also contain an expander such as lactose, sorbitol, sucrose, or mannitol in an amount that facilitates the dispersion of the powder from the device, for example, 50-90% (by weight) of the composition. The preferred enantiomer of 2-hydroxycarboxylic acid should be most advantageously prepared in particle form having an average particle size of less than 10 microns, for example, 0.5-5 microns, for the most effective delivery to the distal lung.

[0263] In one embodiment, the compositions of the disclosed herein may include preservatives, suspending agents, wetting agents, isotonic agents and / or diluents. The compositions provided herein may contain about 0.01% to about 90%, or about 0.01% to about 50%, or about 0.01% to about 25%, or about 0.01% to about 10%, or about 0.01% to about 5%, of a pharmacokinetically suitable suspension of one or more pharmaceutically acceptable amounts when administered by inhalation. Pharmacokinetically suitable fluids for use herein include, but are not limited to, polar solvents (compounds containing hydroxyl groups or other polar groups). Solvents include, but are not limited to, water or alcohols, such as ethanol, isopropanol, and glycols, including propylene glycol, polyethylene glycol, polypropylene glycol, glycol ethers, glycerol, and polyoxyethylene alcohol. Polar solvents also include, but are not limited to, protic solvents, including, water, aqueous saline solutions containing one or more pharmaceutically acceptable salts, alcohols, glycols, or mixtures thereof. In one alternative embodiment, the water used in the composition should meet or exceed the applicable regulatory requirements for use in inhaled medications.

[0264] In one embodiment, the compositions described herein can be aqueous and can contain from 0 to 90% water. In other embodiments, the aqueous compositions described herein can contain from 20 to 80% water. In still other embodiments, the aqueous composition can contain from 50 to 70% water. The water may further include fresh water, distilled water, sterilized water, demineralized water or deionized water. Alternatively, the composition can be non-aqueous and can contain no water or an amount of water that is negligible (e.g., less than 1%, less than 0.1%, less than 0.01%).

[0265] The compositions of the present disclosure may further contain adjuvants such as bronchodilators, anti-inflammatory agents, surfactants, aspirin or ethyl alcohol.

[0266] Bronchodilators optionally used in the compositions of the present disclosure include, but are not limited to, β2-adrenergic receptor agonists (e.g., albuterol, bambuterol, salbutamol, salmeterol, formoterol, arformoterol, levosalbutamol, procaterol, indacaterol, carmoterol, milbeterol, procaterol, terbutaline, etc.) and muscarinic antagonists (e.g., trospium, ipratropium, glycopyrronium, acridinium, etc.). Combinations of drugs may also be used.

[0267] Anti-inflammatory agents that may be used as needed in the compositions of this disclosure include, but are not limited to, inhaled corticosteroids (e.g., beclomethasone, budesonide, ciclesonide, fluticasone, etiprednol, mometasone), leukotriene receptor antagonists and leukotriene synthesis inhibitors (e.g., montelukast, zillotone, ibudilast, zafirlukast, pranlukast, amervant, tiperkast), and cyclooxygenase inhibitors (e.g., ibuprofen, ketoprofen, ketrolac, indomethacin, naproxen, zaltoprofen, lornoxicam, meloxicam, celecoxib, lumiracoxib, etoricoxib, piroxicam, ampiroxicam, cinnoxicam, diclofenac, felbinac, lornoxicam, mesalazine, trifludal, tinoridine, iguratimod, pamicogrel). A combination of drugs may be used. Aspirin may be added to act as an anti-inflammatory agent.

[0268] The surfactants included in this disclosure include, but are not limited to, synthetic surfactants (Exosurf®), dipalmitoylphosphatidylcholine, and oleic acid. Combinations of drugs may also be used. Antioxidants such as glutathione and vitamin E, zinc, and zinc salts of EDTA may be added.

[0269] Ethyl alcohol vapor can act as an antifoaming agent in the lungs, making sputum more liquid, aiding respiration, and reducing pulmonary edema. Ethanol can be added to the compositions of this disclosure in concentrations of 0.5% to 60%, for example, 1 to 40%, 1 to 20%, or 1 to 10%. Ethanol can be added in concentrations of 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, or 60%.

[0270] This disclosure also relates to the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with other drugs delivered by inhalation. These other drugs may include nucleotide sequences that can be incorporated into suitable delivery vectors such as plasmids or viral vectors.Other drugs include therapeutic nucleotide sequences (DNA, RNA, siRNA), enzymes that reduce the viscoelasticity of mucus, such as DNase and other mucolytics, chemicals that upmodulate chloride ion channels or increase the flow of ions across cells, nicotine, P2Y2 agonists, elastase inhibitors, such as α-1 antitrypsin (AAT), N-acetylcysteine, antibiotics and cationic peptides, such as lanthobiotics, specifically duramycin, and short-acting bronchodilators (e.g., albuterol or indacaterol). Examples of anti-inflammatory drugs include β2-adrenergic receptor agonists, M3 muscarinic antagonists (e.g., ipatropium bromide), K channel openers, long-acting bronchodilators (e.g., formoterol, salmeterol), steroids (e.g., budesonide, fluticasone, triamcinolone, beclomethasone, ciclesonide, etc.), xanthines, leukotriene antagonists (e.g., montelukast sodium), phosphodiesterase-4 inhibitors, adenosine receptor antagonists, and various other anti-inflammatory drugs (e.g., Syk kinase inhibitors). (AVE-0950), tryptase inhibitors (AVE-8923 and AVE-5638), tachykinin antagonists (AVE-5883), inducible nitric oxide synthase inhibitors (GW-274150), etc.), transcription factor decoys, TLR-9 agonists, antisense oligonucleotides, siRNA, DNA, CGRP, lidocaine, reverse β2-agonists, anti-infective oxidative therapy, cytokine modulators (e.g., CCR3 receptor antagonists (GSK-766994, DPC-168, AZD-3778), TNF-α production inhibitors (LMP-) These may include small molecule inhibitors of IgE (160 and YS-TH2), IL-4 antagonists (AVE-0309), small molecule inhibitors of IgE, cell adhesion molecule (CAM) inhibitors, small molecules targeting the VLA4 receptor or integrin α4β1 (e.g., R-411, PS-460644, DW-908e, and CDP-323), immunomodulators including those that block T cell signaling by inhibiting calcineurin (tacrolimus), heparin antagonists (talactoferrin α), cytoplasmic PLA2 inhibitors (Efipladib), or combinations thereof.If the subjects requiring CF have CF, they may also be administered in combination with the compositions of this disclosure, standard pharmaceuticals, such as ibacaftol, pulmozyme, mannitol, or other approved drugs according to standard practice.

[0271] Topical delivery excipients Compositions of the present disclosure comprising preferred enantiomers of 2-hydroxycarboxylic acid can be delivered to topical surfaces. For example, topical formulations may be for topical application to the skin, mucous membranes (oral, nasal, vaginal, rectal) or eyes of a target.

[0272] The compositions of this disclosure may contain water (aqueous) or may not contain water.

[0273] The compositions of this disclosure may also contain small amounts of conventional additives, such as viscosity modifiers, such as xanthan gum, and preservatives, such as phenoxyethanol or benzyl alcohol (including mixtures thereof). Some therapeutic agents may require the incorporation of buffers to maintain an appropriate pH.

[0274] Suitable preservatives for use in such compositions or pharmaceuticals include, for example, phenoxyethanol and other preservatives conventionally used in pharmaceutical formulations (especially creams). Suitable preservatives include methyl hydroxybenzoate, chlorocresol, sorbic acid, and benzoic acid.

[0275] The compositions of this disclosure can be manufactured by conventional pharmaceutical techniques. Thus, ointments and creams are conveniently prepared by mixing the components constituting the vehicle together at a high temperature, preferably 60-70°C, until an emulsion is formed. The mixture is then cooled to room temperature, and a preferred enantiomer of 2-hydroxycarboxylic acid is added along with any other components, after which it may be stirred to ensure proper dispersion.

[0276] Liquid preparations such as nasal sprays and eye drops are manufactured by dissolving the therapeutic agent in the components of the vehicle and then adding other components. The resulting solution or suspension is dispensed into glass or plastic bottles, or into single-dose packs such as soft gelatin capsules, and then heat-sealed.

[0277] Artificial tear vehicles can be used in ophthalmic compositions containing preferred enantiomers of 2-hydroxycarboxylic acid delivery. Higher viscosity artificial tears can be produced using high concentrations of thickeners, such as Celluvisc®, high viscosity carboxymethylcellulose (CMC), and Refresh Liquigel®, blends of 0.35% high viscosity CMC and 0.65% low viscosity CMC.

[0278] Gelling agents can be used in compositions containing preferred enantiomers of 2-hydroxycarboxylic acids to be delivered to the eye. Such agents can be dispensed dropwise as a liquid and then almost immediately induce a gel phase. Timoptic gel (gellan gum), AzaSite® (polycarbophil, poloxamer), and Besivance® (polycarbophil, poloxamer), 0.3% alginate Keltrol® are examples of such agents. Another gelling agent is polycarbophil poloxamer gel (e.g., Durasite®).

[0279] The ophthalmic carrier is, in various embodiments, a solvent or dispersion medium containing, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), suitable mixtures thereof, and vegetable oils. Adequate fluidity is maintained, for example, by the use of coatings such as lecithin, control of the required particle size in the case of dispersions, and the use of surfactants. Long-term absorption of the injectable composition is, in certain embodiments, achieved by the use of absorption-delaying agents, such as aluminum monostearate and gelatin in the composition.

[0280] Nasal delivery of the preferred enantiomer of 2-hydroxycarboxylic acid is also intended. Nasal delivery allows the preferred enantiomer of 2-hydroxycarboxylic acid to pass into the bloodstream immediately after administration of the therapeutic product via the nose, without requiring deposition of the preferred enantiomer of 2-hydroxycarboxylic acid in the lungs. Compositions for nasal delivery include those containing dextran or cyclodextran.

[0281] Oral delivery excipients Compositions of the present disclosure comprising preferred enantiomers of 2-hydroxycarboxylic acid can be delivered orally. For example, oral formulations may be for topical or systemic administration to specific parts of the gastrointestinal system (such as the esophagus, stomach, large intestine, or small intestine) and may be delivered in a multitude of forms, including simple solutions, syrups, suspensions, tablets, or capsules.

[0282] The compositions of this disclosure may contain water (aqueous) or may not contain water.

[0283] The compositions of this disclosure may also contain small amounts of conventional additives, such as preservatives, solubilizers (including ethanol), complexing agents (such as cyclodextrin), flowability enhancers and lubricants (including colloidal silica), flavor enhancers and compressibility enhancers (including mixtures thereof). In some therapeutic agents, it may be necessary to incorporate buffers to maintain an appropriate pH.

[0284] The oral composition of the present invention may be delivered together with an immunostimulant or modifier. Examples of immunostimulants or modifiers include specific immunostimulants (such as vaccines and antigens) and nonspecific agents, such as bee products (including propolis and honey), probiotics and prebiotics, hormones, vitamins (vitamin C, vitamin D), minerals (zinc oxide), antioxidants (including glutathione), interferons (including INF-alpha), interleukins (including interleukin-10), colchicine, thalidomide, and imiquimod. The immune booster may be delivered simultaneously with or after the preferred enantiomer of the 2-hydroxycarboxylic acid composition.

[0285] In some situations, administration of a preferred enantiomer of 2-hydroxycarboxylic acid may sufficiently dysregulate the metabolism of planktonic bacteria in the body, and as a result, it is thought that the body may be able to eliminate planktonic bacteria without the use of further antimicrobial agents such as antibiotics, either concurrently or subsequently with immunostimulants.

[0286] Plant delivery excipients This composition may contain active pesticide compounds other than the preferred enantiomer of the 2-hydroxycarboxylic acid of the present invention.

[0287] This composition may further contain one or more of the following auxiliary components: inert carriers, surfactants such as adhesives or spreading agents, stabilizers and / or dyes, etc. This composition may also be suspended in a carrier fluid such as air, nitrogen, carbon dioxide or fumigation gas. This composition preferably contains auxiliary agents, such as fillers, solvents, spontaneous action enhancers, carriers, emulsifiers, dispersants, cryoprotectants, biocides, thickeners and / or other auxiliary agents, such as adjuvants. An adjuvant in this context is a component that enhances the biological effect of the composition without having a biological effect itself. An example of an adjuvant is an agent that promotes the retention, diffusion, adhesion to leaf surfaces, or penetration of a preferred enantiomer of a 2-hydroxycarboxylic acid into plant tissue. Generally speaking, the active compound can be combined with any solid or liquid additive commonly used for formulation purposes.

[0288] The compositions of the present invention may also include formulation aids and additives known to those skilled in the art as formulation aids (some of which may also function as solid diluents, liquid diluents, or surfactants). Such formulation aids and additives can control pH (buffering agents), foaming during processing (defoaming agents, e.g., polyorganosiloxanes), sedimentation of active ingredients (suspending agents), viscosity (thixotropic or pseudoplastic thickeners), growth of microorganisms in containers (antimicrobial agents), freezing of the product (antifreeze agents), coloring (dye / pigment dispersions), washability (film-forming agents or adhesives), evaporation (evaporation retarders), and other formulation properties. Examples of film-forming agents include polyvinyl acetate, polyvinyl acetate copolymers, polyvinylpyrrolidone-vinyl acetate copolymers, polyvinyl alcohol, polyvinyl alcohol copolymers, and waxes.

[0289] stability Where necessary, the compositions of this disclosure are stable. When used herein, the stability of the compositions provided herein refers to the length of time that more than 80%, 85%, 90%, or 95% of the initial amount of antibiotic-lactic acid is present in the composition at a given temperature. For example, the compositions provided herein may be stored at about 15°C to about 30°C and remain stable for at least 1, 2, 12, 18, 24, or 36 months. Alternatively, the compositions may be suitable for administration to subjects requiring them after being stored at 25°C for longer than 1, 2, 12, 18, 24, or 36 months. In another alternative embodiment, Arrhenius dynamics are used to show that more than 80%, or more than 85%, or more than 90%, or more than 95% of the initial amount of active substance (e.g., a preferred enantiomer of 2-hydroxycarboxylic acid) remains after the composition has been stored at about 15°C to about 30°C for longer than 1, 2, 12, 18, 24, or 36 months.

[0290] Where used herein, the statement that a composition is stable during “long-term storage” means that the composition is suitable for administration to subjects requiring it if it has an estimated shelf life of more than one, two, or three months of use at 25°C and more than one, two, or three years of storage at 5°C. In certain embodiments herein, Arrhenius dynamics are used to determine that an estimated >80% or >85% or >90% or >95% of the preferred enantiomers of the 2-hydroxycarboxylic acid remain after such storage.

[0291] Antibiotics Other activators may also be incorporated into the compositions of this disclosure. For example, additional antimicrobial compounds such as antibacterial agents and antifungal agents may be incorporated.

[0292] Additional activators provided with the preferred enantiomers of the 2-hydroxycarboxylic acids of this disclosure may be bacterial-killing activators, but are not antibiotics. For example, additional activators may be chlorine-based compounds (such as bleaches), iodine-based compounds, copper-based compounds, and the like.

[0293] The preferred enantiomers of the 2-hydroxycarboxylic acid of this disclosure are preferably dysregulated in the metabolism of planktonic bacteria so that an additional activator (antibiotic or other compound capable of killing bacteria) can have a better bactericidal effect. If necessary, the metabolism of the planktonic bacteria is dysregulated to such an extent that the bacteria cannot resist the effects of the additional activator. The preferred enantiomers of the 2-hydroxycarboxylic acid of this disclosure are even more preferably dysregulated in the metabolism of planktonic bacteria so that the planktonic bacteria do not develop resistance to the additional activator.

[0294] Metabolic dysregulation in planktonic bacteria in the presence of the preferred enantiomer of 2-hydroxycarboxylic acid alone may result in either suppression or enhancement of the bacterial metabolism. Following the introduction of an antimicrobial compound (so that the bacteria are exposed to both the antimicrobial compound and 2-hydroxycarboxylic acid), the dysregulation (suppression or enhancement) may persist, possibly with an increased degree of dysregulation. However, when planktonic bacteria are present with both the preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, the metabolic dysregulation of the planktonic bacteria may change from previously suppressive to enhancement, and from previously enhanced to suppression. Alternatively, metabolic dysregulation may not occur in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation may occur in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and an antimicrobial compound, particularly if the planktonic bacteria were resistant to the antimicrobial compound in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid.

[0295] The composition may further contain benzoyl peroxide, or an antibiotic such as erythromycin, clindamycin, doxycycline, or meclocycline.

[0296] Additional antimicrobial compounds that can be used include, but are not limited to, silver compounds (e.g., silver chloride, silver nitrate, silver oxide), silver ions, silver particles, iodine, povidone / iodine, chlorhexidine, 2-p-sulfanylanilinoethanol, 4,4'-sulfinyldianiline, 4-sulfanilamide salicylic acid, acediasulfone, acetosulfone, amikacin, amoxicillin, amphotericin B, ampicillin, aparcillin, apicycline, apramycin, arbekacin, aspaxicillin, azidanphenicol, azithromycin, and aztreo. Nam, bacitracin, vanbelmycin (multiple possible), biapenem, brodimoprim, butyrosine, capreomycin, carbenicillin, carbomycin, carmonam, cefadroxil, cephamandol, cefatoridine, cefbuperazone, cefclizine, cefdinir, cefditoren, cefepime, cefetamet, cefixime, cefinenoxime, cefminox, cefozidime, cefonicid, cefoperazone, cefolanide, cefotaxime, cefotetan, cefotian, cefozopran, cefpimisole, cefpyramide, cefpirome, cef Fuprodil, cefuroxazine, ceftazidime, ceferam, ceftibutene, ceftriaxone, cefzonam, cephalexin, cephaloglysin, cephalosporin C, cefaflavine, chloramphenicol, chlortetracycline, ciprofloxacin, clarithromycin, clinafloxacin, clindamycin, chromocycline, colistin, cyclacillin, dapsone, demeclocycline, diatimosulfone, dibekacin, dihydrostreptomycin, dilithromycin, doxycycline, enoxacin, enviomycin, episi Phosphorus, erythromycin, flomoxef, fortimycin (multiple), gentamicin (multiple), glucosulfone, solasulfone, gramicidin S, gramicidin (multiple), glepafloxacin, guamecycline, hetacillin, imipenem, isepamycin, josamycin, kanamycin (multiple), leucomycin (multiple), lincomycin, lomefloxacin, lusimycin, rimescycline, meclocycline, meropenem, metacycline, micronomycin, midecamycin (multiple), minocycline, moxalactam, mupirocin,Nadifloxacin, natamycin, neomycin, netylmycin, norfloxacin, oleandmycin, oxytetracycline, p-sulfanylbenzylamine, panipenem, paromomycin, pazufloxacin, penicillin N, pipecycline, pipemidic acid, polymyxin, primycin, quinacillin, ribostamycin, rifamide, rifampin, rifamycin SV, rifapentin, rifaximin, ristocetin, litipenem, rokitamycin, lolitetracycline, rosalamycin, roxithromycin, sulfasalazine, sancycline, shisomycin, sparfloxacin, spectinomycin, spiramycin, streptomycin, succisulfone, sulfacylisoidine, sulfaloxine, sulfamidequaidine, sulfani Examples include temafloxacin, sulfoxone, teicoplanin, temafloxacin, temocillin, tetracycline, tetroxoprim, thiamphenicol, thiazol sulfone, thiostrepton, ticarcillin, tigemonam, tobramycin, tosufloxacin, trimethoprim, trospectomycin, trovafloxacin, tubeactinomycin, vancomycin, azaserin, candicidine (multiple), chlorphenesin, dermostatin (multiple), philipin, fundicromin, mepaltricin, nystatin, oligomycin (multiple), ciproflaxacin, norfloxacin, ofloxacin, pefloxacin, enoxacin, losofloxacin, amifloxacin, freloxacin, temafloxacin, lomefloxacin, perimycin A, or tubercidine.

[0297] use This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the preparation of compositions for dysregulating the metabolism of planktonic bacteria.

[0298] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for metabolic dysregulation in planktonic bacteria.

[0299] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the preparation of compositions for dysregulating the metabolism of planktonic bacteria in combination with antimicrobial compounds.

[0300] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds to dysregulate the metabolism of planktonic bacteria.

[0301] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in the preparation of compositions for treating or preventing infections in a subject, wherein the infection is caused by planktonic bacteria, and the composition dysregulates the metabolism of the planktonic bacteria.

[0302] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds in the preparation of compositions for treating or preventing infections in a subject, wherein the infection is caused by planktonic bacteria, and the composition dysregulates the metabolism of the planktonic bacteria.

[0303] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids for treating or preventing infections in a subject, wherein the infection is caused by planktonic bacteria, and the composition dysregulates the metabolism of the planktonic bacteria.

[0304] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acids in combination with antimicrobial compounds to treat or prevent infections in a subject, wherein the infection is caused by planktonic bacteria, and the composition dysregulates the metabolism of the planktonic bacteria.

[0305] This disclosure provides the use of preferred enantiomers of 2-hydroxycarboxylic acid in the preparation of compositions for sensitizing planktonic bacteria to antimicrobial compounds, wherein preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0306] The disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acid to sensitize planktonic bacteria to antimicrobial compounds, wherein the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in planktonic bacteria.

[0307] This disclosure provides the use of a preferred enantiomer of 2-hydroxycarboxylic acid in the preparation of a composition for sensitizing planktonic bacteria to an antimicrobial compound, wherein the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0308] The disclosure further provides the use of a preferred enantiomer of 2-hydroxycarboxylic acid to sensitize planktonic bacteria to an antimicrobial compound, wherein the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in the planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0309] The disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acid in the preparation of compositions for treating or preventing infections in a subject, wherein the infection is caused by planktonic bacteria, the composition sensitizes the planktonic bacteria to an antimicrobial compound, and the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacteria.

[0310] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acid in the preparation of compositions for treating or preventing infections in a subject, in combination with antimicrobial compounds, wherein the infection is caused by planktonic bacteria, and the composition sensitizes the planktonic bacteria to the antimicrobial compound, and the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacteria.

[0311] This disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acid for treating or preventing an infection in a subject, wherein the infection is caused by planktonic bacteria, and the composition sensitizes the planktonic bacteria to the antimicrobial compound, with the preferred enantiomers of 2-hydroxycarboxylic acid inducing metabolic dysregulation in the planktonic bacteria.

[0312] The disclosure further provides the use of preferred enantiomers of 2-hydroxycarboxylic acid in combination with antimicrobial compounds to treat or prevent infections in a subject, wherein the infection is caused by planktonic bacteria, and the composition sensitizes the planktonic bacteria to the antimicrobial compound, and the preferred enantiomers of 2-hydroxycarboxylic acid induce metabolic dysregulation in the planktonic bacteria.

[0313] In the above use, metabolic dysregulation of planktonic bacteria in the presence of the preferred enantiomer of 2-hydroxycarboxylic acid alone may be either suppression or enhancement of the planktonic bacteria's metabolism. After the introduction of the antimicrobial compound, the dysregulation (suppression or enhancement) may persist, possibly with an increase in the degree of dysregulation. However, when planktonic bacteria are in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, metabolic dysregulation of the planktonic bacteria may change from previously suppressive to enhancement, and from previously enhanced to suppression. Alternatively, metabolic dysregulation may not occur in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation may occur in the presence of the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, especially if the planktonic bacteria were resistant to the antimicrobial compound in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid.

[0314] The preferred enantiomer of the 2-hydroxycarboxylic acid may have the same composition as the antimicrobial compound, or it may have a different composition. The airborne bacteria may be associated with or capable of causing the infection of the target, or they may be present on surfaces such as non-living surfaces. The airborne bacteria may be present in living or non-living liquids or gases.

[0315] If a preferred enantiomer of 2-hydroxycarboxylic acid is present in a composition separate from the antimicrobial compound, the preferred enantiomer of 2-hydroxycarboxylic acid may be administered simultaneously with the antimicrobial compound or at a different time from the antimicrobial compound.

[0316] For the above use, the preferred enantiomer of 2-hydroxycarboxylic acid may be D-lactic acid. If necessary, infection or colonization is bacterial infection or colonization.

[0317] kit This disclosure provides a kit for dysregulating the metabolism of planktonic bacteria, and this kit is, a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes the instruction manual.

[0318] This disclosure provides a kit for dysregulating the metabolism of planktonic bacteria, and this kit is, a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes instructions for use, Here, a preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0319] This disclosure provides a kit for sensitizing planktonic bacteria to an antimicrobial compound, and this kit is a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes instructions for use, Here, the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria.

[0320] This disclosure provides a kit for sensitizing planktonic bacteria to an antimicrobial compound, and this kit is a) Preferred enantiomers of 2-hydroxycarboxylic acid, b) Includes instructions for use, Here, the preferred enantiomer of 2-hydroxycarboxylic acid induces metabolic dysregulation in planktonic bacteria, and the preferred enantiomer of 2-hydroxycarboxylic acid is combined with an antimicrobial compound.

[0321] In the above kit, metabolic dysregulation of planktonic bacteria in the presence of the preferred enantiomer of 2-hydroxycarboxylic acid alone may result in either suppression or enhancement of the planktonic bacteria's metabolism. After the introduction of the antimicrobial compound, the dysregulation (suppression or enhancement) may persist, possibly with an increased degree of dysregulation. However, when planktonic bacteria are present with both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, metabolic dysregulation of the planktonic bacteria may change from previously suppressive to enhanced, and from previously enhanced to suppressive. Alternatively, metabolic dysregulation may not occur in the presence of 2-hydroxycarboxylic acid alone, but significant dysregulation may occur in the presence of both the preferred enantiomer of 2-hydroxycarboxylic acid and the antimicrobial compound, especially if the planktonic bacteria were resistant to the antimicrobial compound in the absence of the preferred enantiomer of 2-hydroxycarboxylic acid.

[0322] A preferred enantiomer of the 2-hydroxycarboxylic acid in the kit may be D-lactic acid.

[0323] If the components of the kit are provided in one or more liquid solutions, the liquid solutions may be aqueous solutions, such as sterile aqueous solutions. For in vivo use, the expression construct may be formulated into a pharmaceutically acceptable injectable composition. In this case, the container means may itself be an inhaler, syringe, pipette, eye dropper, or other such device from which the composition can be applied to a diseased area of ​​the animal, such as the lungs, injected into the animal, or applied to and even mixed with other components of the kit.

[0324] In one embodiment, the kit of the present disclosure comprises a composition comprising a preferred enantiomer of a 2-hydroxycarboxylic acid, optionally combined with an antimicrobial compound. In an alternative embodiment, the composition is pre-measured, pre-mixed, and / or pre-packaged.

[0325] The kits of the present disclosure may also include instructions designed to facilitate user compliance. As used herein, instructions refer to any labels, inserts, etc., which may be placed on one or more surfaces of the packaging material, or the instructions may be provided on separate sheets or any combination thereof. For example, in one embodiment, the kit of the present disclosure includes instructions for administering the composition of the present disclosure. In one embodiment, the instructions indicate that the composition of the present disclosure is suitable for metabolic dysregulation of planktonic bacteria. Such instructions may also include instructions for dosage and instructions for administration.

[0326] Preferred enantiomers of 2-hydroxycarboxylic acids and suitable excipients can be packaged individually so that the practitioner or user can formulate the components into pharmaceutically acceptable compositions as needed. Alternatively, the antisense oligomer and suitable excipients can be packaged together, thereby requiring minimal composition by the practitioner or user. In either case, the packaging should maintain the chemical, physical, and aesthetic integrity of the active ingredient.

[0327] general Those skilled in the art will understand that the present invention as described herein is susceptible to modifications and alterations other than those specifically described. The present invention includes all such modifications and alterations. The present invention also includes all of the steps, features, compositions and compounds mentioned or indicated herein, individually or collectively, and in any combination or of any two or more steps or features.

[0328] Each document, reference, patent application, or patent cited herein is expressly incorporated in its entirety by reference, meaning that it should be read and considered by the reader as part of this document. The absence of repetition of any document, reference, patent application, or patent cited herein is solely for the sake of brevity.

[0329] Any manufacturer's instructions, descriptions, product specifications, and product sheets relating to any product mentioned herein or in any document incorporated herein by reference may be incorporated herein by reference and used in the practice of the present invention.

[0330] The scope of the present invention is not limited by any of the specific embodiments described herein. These embodiments are for illustrative purposes only. Functionally equivalent products, compositions, and methods are clearly within the scope of the present invention as described herein.

[0331] The inventions described herein may include ranges of values ​​greater than 1 (e.g., size, displacement, and electric field strength). A range of values ​​is understood to include all values ​​within the range, including the value that defines the range and the values ​​adjacent to the range that produce the same or substantially the same results as the values ​​directly adjacent to the value that defines the boundary to the range. Thus, unless otherwise indicated, the numerical parameters described herein and in the claims are approximations that may vary depending on the desired properties to be obtained by the invention. Thus, “about 80%” means “about 80%” and also “80%”. At a minimum, each numerical parameter should be interpreted in light of the number of significant figures and common rounding techniques.

[0332] Throughout this specification, unless otherwise required by context, the word “comprise,” or variations such as “comprises” or “comprising,” shall be understood to mean that they include the integers or sets of integers described, but do not exclude any other integers or sets of integers. Furthermore, it should be noted that in this disclosure, particularly in the claims and / or paragraphs, terms such as “comprises,” “comprised,” and “comprising” may have meanings attributable to them under U.S. patent law. For example, they may mean “includes,” “included,” and “including.” Terms such as “consisting essentially of” and “consists essentially of” have meanings attributable to them under U.S. patent law, for example, they allow elements not explicitly enumerated but exclude elements found in the prior art or that affect the fundamental or novel features of the present invention.

[0333] Other definitions of selected terms used herein are found in the detailed description of the invention and may apply throughout. Unless otherwise defined, all other scientific and technical terms used herein have the same meaning as commonly understood by those skilled in the art to which the invention pertains. The term “activator” may mean one activator, or may encompass two or more activators.

[0334] The following examples will more fully illustrate the methods of using the present invention as described above, and will help to illustrate the best modes intended for carrying out various aspects of the invention. It will be understood that these methods are not in any way limited to the true scope of the invention, but rather are presented for illustrative purposes. [Examples]

[0335] Examples

[0336] Further features of the present invention are described more fully in the following non-limiting embodiments. This description is included solely for illustrative purposes and should not be understood as a limitation to the broader description of the invention described above.

[0337] Example 1 Suppression of the metabolism of planktonic bacteria This study demonstrates that D-lactic acid inhibits the metabolism of planktonic bacteria.

[0338] material and method Floating bacterial culture Clinical isolates of Pseudomonas aeruginosa collected from patients with cystic fibrosis were phenotypically classified and stored as 50% glycerol stocks at -70°C ± 2°C. The clinical isolates were thawed and streaked onto blood agar plates, where they were grown at 37°C, 5% CO2 for 12–24 hours, or until the isolated colonies were large enough to sustain further culture and could be selected. The isolated colonies were inoculated in a loop into 5 mL of LB broth and incubated overnight at 37°C, 5% CO2 with gentle shaking until the stationary phase (B0) was reached.

[0339] Dilute the B0 culture in Pseudomonas Minimal Medium (referred to as PMM in this specification) to a ratio of 1:1000 and prepare for approximately 10 8 After achieving cfu / mL, 150 μL of each diluted culture was transferred to the wells of a 96-well microtiter tray and incubated overnight at 37°C, 5% CO2. D-lactate was then applied by adding 50 μL of appropriate stock solution diluted with PMM (or vehicle only in the case of control wells) and incubating for a further 24 hours at 37°C, 5% CO2. Next, 20 μL of the culture supernatant was transferred to a well containing 180 μL of fresh PMM, and the culture density was determined by measuring the absorbance at 595 nm (using a ThermoFisher Multiskan FC plate reader). Metabolic activity was then determined using these diluted cultures by resazurin staining.

[0340] Resazurin dyeing Changes in bacterial metabolic activity after treatment were measured using resazurin staining (Kirchner et al., 2012). 10 μl of 0.02% (v / v) resazurin (diluted with distilled water) was added to each well, and the microtight plate was incubated under aerobic conditions at 37°C for 1–2 hours with shaking at 150 rpm. Live cells reduce the blue resazurin dye to a pink fluorescent resorphine form. After incubation, fluorescence was quantified using an Enspire microplate reader with an excitation wavelength of 540 nm and an emission wavelength of 590 nm.

[0341] statistical analysis All data presented represent data from 24 clinical isolates of Pseudomonas aeruginosa, each containing two technical copies, unless otherwise specified. Student's t-test was used to assess significance compared to the control untreated biofilm, with significance measured at p<0.05.

[0342] result The effect of D-lactate on the metabolic activity of suspension bacterial cultures was quantified by measuring resolphin fluorescence. In cultures treated with D-lactate, 90% inhibition of metabolic activity was observed, and the change in activity was most pronounced at D-lactate concentrations between 0.25 mg / mL and 0.5 mg / mL (Figure 1).

[0343] D-lactic acid does not significantly reduce the floating biomass of P. aeruginosa, but it has a significant effect on baseline metabolic activity.

[0344] Example 2 Inhibiting the metabolism of planktonic bacteria increases microbial susceptibility to antimicrobial compounds. This study demonstrates that D-lactic acid treatment increases the susceptibility of planktonic bacteria to antibiotics to such an extent that antimicrobial resistance becomes reversible with D-lactic acid treatment.

[0345] material and method Floating bacterial culture Bacterial strains were maintained at -80°C as suspensions in 20% (v / v) glycerol / brainheart infusion. For use, loop-full ice was scraped from the stock, streaked onto nutrient-rich medium containing 1.8% agar to obtain single colonies, and incubated overnight at 37°C (Luria-Bertani medium was used for Pseudomonas aeruginosa or Klebsiella pneumoniae, and triptycase soy agar for Staphylococcus aureus or Streptococcus pneumoniae).

[0346] Minimum inhibitory concentration by broth microdilution method Standard inoculum were prepared from broth cultures. One to three colonies from an overnight agar culture plate were transferred to 5 ml of broth medium and incubated at 37°C (200 rpm) with shaking until the turbidity was greater than the 0.5 McFarland standard (usually overnight). The following broth media were used: cation-modified Mueller-Hinton broth (caMHB - whole strain); Roswell Park Memorial Laboratory medium (RPMI1640, phenol red-free - S. aureus); triptycase Sawyer broth (TSB - S. aureus, S. pneumoniae); Todd-Hewitt broth (THB - S. pneumoniae); M63 minimal medium supplemented with 0.2% glucose (M63 - K. pneumoniae).

[0347] The broth culture was diluted in broth to a turbidity equal to that of a 0.5 McFarland standard, measured using a spectrophotometer at a wavelength of 620 nm. The 0.5 McFarland standard was further diluted 1:100 in broth to prepare a standard inoculum. 50 μl of this standard inoculum was dispensed into all wells of a microtiter tray, while 50 μl of broth was dispensed instead for the negative control.

[0348] Next, for all antimicrobial agents, a working concentration twice the highest target concentration was prepared from the stock solution in broth. 100 μl of this working concentration was pipetteed into the first well, and 50 μl of broth medium was added to all other wells. 50 μl of the first well was pipettered and serially diluted across all wells in that row until the final concentration within the range was achieved. 50 μl was taken from the final well and discarded. This resulted in a concentration range where each well contained twice the target concentration. This was performed in double doses for all agents.

[0349] 96-well trays were incubated at 37°C for 18 ± 2 hours. After incubation, the MIC was visually assessed and recorded as the lowest concentration of antimicrobial agent that completely inhibited bacterial growth, resulting in a complete absence of turbidity.

[0350] Resazurin reduction for evaluation of metabolic activity Aerobic respiration was measured after incubation and visual recording of MIC, or in a separate study, in the presence or absence of D-lactic acid and concomitant antibiotics. For evaluation after completion of MIC determination, the entire volume (100 μl) of the MIC wells was mixed by pipetting up and down and then transferred to the corresponding wells of a 96-well plate suitable for fluorescence and absorbance measurements. Alternatively, the cultures were pipetteed together as described above and incubated overnight at 37°C in the absence of antimicrobial compounds. The culture supernatant was removed, adherent cells were washed, and then treated by adding 100 μL of fresh broth, 50 μL of broth containing 4x the desired concentration of antibiotic, and 50 μL of broth containing 4x the desired concentration of D-lactic acid or vehicle only as a control group, and then incubated for a further 18 ± 2 hours at 37°C. After the incubation regime was complete for both procedures, resazurin reduction was evaluated by adding 10 μl of 0.02% resazurin solution to each well and incubating for a further 90 minutes at 37°C with gentle shaking of the plate (40 rpm). The 96-well plate and resazurin were protected from light as much as possible.

[0351] After incubation, the 96-well tray was read using a fluorescence absorbance spectrophotometer with an excitation wavelength of 565 nm and an emission wavelength of 590 nm. Blank correction was performed on the values ​​of each well by subtracting the values ​​of the negative control. Then, all values ​​were converted to ratios to the mean values ​​of the positive control.

[0352] result The minimum inhibitory concentration (MIC) is reduced by the simultaneous administration of D-lactate with multiple antibiotics. First, the MICs of multiple antibiotics, either alone or in combination with D-lactic acid, were determined for medically relevant strain / antibiotic combinations. The results are shown in Table 3, indicating that D-lactic acid treatment resulted in at least a twofold reduction in MIC, i.e., at least a twofold increase in the potency of the antibiotic against the bacterial strain. [Table 3] The inhibition of the metabolism of planktonic bacteria by D-lactic acid is further enhanced by antibiotic treatment.

[0353] The effect of D-lactic acid on the metabolic activity of suspension cultures when combined with either a bactericidal or bacteriostatic antibiotic was quantified by measuring resolphin fluorescence. The data are summarized in Table 4 and shown in Figures 2 to 10. [Table 4]

[0354] The decrease in metabolic activity observed during antibiotic treatment was further enhanced when combined with D-lactate. In Figures 2–10, the histograms show the amount of metabolic activity measured by resorphine fluorescence compared to a vehicle-only control. Relative metabolic activity is shown in black for antibiotic-only treatment and in gray for antibiotic + D-lactate treatment.

[0355] Example 3 Metabolic dysregulation in planktonic bacteria is specific to D-enantiomers. This study demonstrates that the metabolic dysregulation observed in P. aeruginosa (Pseudomonas aeruginosa) in the presence of D-lactate is not reproduced in the presence of L-lactate.

[0356] material and method Floating bacterial culture P. aeruginosa strain WACC91 was maintained at -80°C as a suspension in 20% (v / v) glycerol / brainheart infusion. For use, a loop full of ice was scraped from the stock and streaked for single colonies on LB medium containing 1.8% agar, and incubated overnight at 37°C. Isolated colonies were placed by loop into 5 mL of LB broth and incubated at 37°C with shaking until turbidity was greater than 1.0 McFarland standard (3-4 hours).

[0357] The broth culture was diluted in M63 minimal medium containing 0.2% (w / v) glucose as the sole carbon source to a turbidity equal to 1.0 McFarland standard at a wavelength of 595 nm using a spectrophotometer. This culture was then diluted 1:100 with M63 (glucose), and 100 μL, along with an additional 50 μL, was transferred to each well of a 96-well microtiter tray and incubated overnight at 37°C, 5% CO2. D-lactic acid, L-lactic acid, or vehicle alone (sterile H2O) was then applied by adding 50 μL of a suitable stock solution diluted with M63 (glucose) and incubating for a further 24 hours at 37°C, 5% CO2. The culture metabolic activity was then determined using resazurin reduction as described above in Example 1.

[0358] result Resorphin fluorescence was measured to quantify the effects of D-lactate or L-lactate on the metabolic activity of suspension cultures. Cultures treated with D-lactate showed >50% inhibition of metabolic activity (compared to treatment with vehicle alone), and the changes in activity were most pronounced with D-lactate concentrations between 12.5 mM and 25 mM (Figure 1). No changes in metabolic activity were observed in cultures treated with L-lactate. Therefore, the observed metabolic dysregulation is specific to the D-enantiomer of lactate.

[0359] Figure 11 shows histograms of P. aeruginosa WACC91 suspension cultures after the addition of D-lactic acid or L-lactic acid at 2-fold serial dilutions. The figure shows that, compared to treatment with vehicle alone, a breakpoint is observed with D-lactic acid between 12.5 mM and 25 mM, using metabolic rate (reduction of resazurin to fluorescent resorphine) as an indicator. No equivalent breakpoint is observed when L-lactic acid is added at the same concentration at which the effect is observed with D-lactic acid.

[0360] Example 4 D-lactate enhances the antibacterial activity of metal ions. This study demonstrates that the antimicrobial activity of metal ions against planktonic bacteria is enhanced when metal ions are present as D-lactates.

[0361] material and method Floating bacterial culture The bacterial strains were maintained at -80°C as suspensions in 20% (v / v) glycerol / brainheart infusion. For use, a loop full of ice was scraped from the stock, and single colonies were streaked on triptycase soy agar containing 1.8% agar and incubated overnight at 37°C.

[0362] Minimum inhibitory concentration by broth microdilution method Standard inoculum was prepared from broth cultures. 1–3 colonies from an overnight agar plate were transferred to 5 ml of broth medium and incubated overnight at 37°C with shaking at 200 rpm until the turbidity was greater than that of a 0.5 McFarland standard. All assays were performed in triptycase soya broth.

[0363] The broth culture was diluted in broth to a turbidity equal to that of a 0.5 McFarland standard, measured using a spectrophotometer at a wavelength of 620 nm. The 0.5 McFarland standard was further diluted 1:100 in broth to prepare a standard inoculum. 50 μl of this standard inoculum was dispensed into all wells of a microtiter tray, while 50 μl of broth was dispensed instead for the negative control.

[0364] Next, working concentrations twice the maximum target concentration for all salts (in aqueous solution) were prepared in the broth from the stock solution. 100 μl of this working concentration was pipetteed into the first well, and 50 μl of the broth medium was added to all the other wells. 50 μl of the first well was pipettered and serially diluted across all wells in that row until the final concentration within the range was achieved. 50 μl was taken from the final well and discarded. This resulted in a concentration range where each well had twice the target concentration. This was performed in double steps for all agents.

[0365] 96-well trays were incubated at 37°C for 18 ± 2 hours. After incubation, the MIC was visually assessed and recorded as the lowest concentration of antimicrobial agent that completely inhibited bacterial growth, resulting in a complete absence of turbidity.

[0366] result The minimum inhibitory concentration (MIC) decreases in the presence of D-lactic acid. The MICs of zinc (Zn) or copper (Cu) salts present as nitrates or D-lactates were determined. The results are shown in Table 4, demonstrating that the presence of D-lactates results in at least a twofold decrease in MIC, i.e., at least a twofold increase in the potency of the metal ion against the bacterial strain in question. [Table 5]

[0367] Example 6 Inhibiting the metabolism of planktonic bacteria increases the susceptibility of microorganisms to antimicrobial compounds. This study demonstrates that D-lactate treatment enhances the susceptibility of colistin-resistant strains of Escherichia coli to colistin in a mouse peritoneal sepsis model, to the extent that combined D-lactate and colistin treatment, compared to colistin alone, (i) mitigates disease severity as measured by imaging of viable bacterial counts in blood samples and bacterial load in living animals, and (ii) increases mouse survival rate.

[0368] material and method Preparation of bacterial inoculum Bioluminescent col R E. coli Xen14 was streaked onto a 5% sheep blood agar plate containing 30 μg / ml kanamycin and incubated overnight (o / n) at 37°C in normal air. First, the plate was examined using a Lumina XRMS III instrument to confirm bioluminescence, and then one loopful of the overnight culture was mixed into 10 mL of Luria Bertani (LB) broth. 600 The suspension was adjusted to nm=0.05. 600nm=0.5 (approx. 5×10 8 The suspension was incubated at 37°C for approximately 3 hours until it reached a concentration equivalent to CFU / mL. The bacterial suspension was centrifuged at 4,000 × g for 5 minutes, resuspended, washed twice in phosphate-buffered saline (PBS, pH 7.2), and resuspended again in PBS. It was then placed on moist ice and used for mouse infection within 10 minutes of preparation, as described below.

[0369] Mouse infection research These experiments used 6-7 week old (approximately 26-33g, 8 mice per group) uninbred Swiss male (CD1) mice obtained from the Laboratory Animal Services breeding facility at the University of Adelaide. The mice were given free access to food and water. The Animal Ethics Committee at the University of Adelaide (Approval No. S-2022-044) reviewed and approved all animal experiments. This study was conducted in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes (8th Edition 2013) and the South Australian Animal Welfare Act 1985.

[0370] Each mouse was placed in 200 μL PBS containing 3% porcine gastric mucin type III (Sigma Aldrich) for approximately 1 × 10⁶ mice. 8 CFU bioluminescence R E. coli Xen14 was challenged intraperitoneally (IP), and bioluminescence imaging was performed under anesthesia on the IVIS Lumina XRMS Series III system at both ventral and dorsal positions. Immediately afterward, the efficacy of sodium D-lactate (1000 mg / kg; pH 7.03) was tested as follows. group: Group 1: Single oral dose of vehicle-only control (sterile water only) Group 2: Single oral dose of 1000 mg / kg of sodium D-lactate (active ingredient 1) in water. Group 3: Single oral dose of 1000 mg / kg of sodium D-lactate at 0 hours + four IP doses of 1 mg / kg of colistin (col) at 0, 4, 8, and 12 hours (active ingredient 2) Group 4: Four IP doses of 1 mg / kg colistin (col) at 0, 4, 8, and 12 hours (control drug below therapeutic dose).

[0371] Aliquots (approximately 50 μL) of blood samples were collected from each mouse in all groups over a 4-hour period by submandibular (facial vein) blood sampling for bacterial counting. Mice were frequently observed for signs of distress up to 72 hours after infection, and their clinical condition was recorded and imaged. Mice that were near death or showed any evidence of distress were humanely euthanized by CO2 asphyxiation. In all experiments, signals were collected from a defined region of interest, and total flux intensity (photons / second) was analyzed using Living Image Software 4.7.3. Survival rates for each group were recorded. Differences in bioluminescence signals between groups were compared using unpaired t-tests (two-tailed).

[0372] result As detailed above, mice are placed in 200 μL PBS for approximately 1 × 10⁶ 8 CFU bioluminescence R Intraperitoneal (IP) challenge was performed with Escherichia coli Xen14, followed by treatment with one of the following: a single oral dose of sterile water (negative control), a single oral dose of sodium D-lactate (1000 mg / kg), a combination of a single oral dose of sodium D-lactate (1000 mg / kg) and four IP doses of colistin (1 mg / kg at 0, 4, 8, and 12 hours), or four IP doses of colistin (1 mg / kg at 0, 4, 8, and 12 hours) alone.

[0373] The effectiveness of the treatment was evaluated by: the number of colonies in blood samples taken 4 hours after infection; and fluorescently tagged colonies. RImaging of infected mice for E. coli concentration; and survival (Figures 12-14). Mice treated with a single dose of 1000 mg / kg sodium D-lactate and four doses of 1 mg / kg colistin had significantly lower colony counts from blood samples than mice treated with four doses of 1 mg / kg colistin alone. A single dose of 1000 mg / kg sodium D-lactate alone was not protective. Fluorescently tagged col R Similar effects were observed in both imaging and survival rates of infected mice regarding E. coli concentration.

Claims

1. A composition for dysregulating the metabolism of planktonic bacteria, wherein the composition has the following formula: 【Chemistry 4】 A composition comprising 2-hydroxycarboxylic acid.

2. The aforementioned 2-hydroxycarboxylic acid is given by the following formula: 【Transformation 5】 The composition according to claim 1, which is a preferred enantiomer of a 2-hydroxycarboxylic acid having the following characteristics.

3. The following formula: 【Transformation 6】 The composition according to claim 2, further comprising an undesirable enantiomer of a 2-hydroxycarboxylic acid having the above.

4. The composition according to claim 3, wherein the percentage of undesirable enantiomers of 2-hydroxycarboxylic acid is less than 20% of the total amount of 2-hydroxycarboxylic acid.

5. The composition according to claim 2, wherein the composition does not contain an undesirable enantiomer of 2-hydroxycarboxylic acid.

6. The aforementioned dysregulation, (i) Inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid alone, and inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (ii) Inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid alone, but enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (iii) Enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid alone, and enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (iv) Enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid alone, but inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (v) No effect on metabolism in the presence of the 2-hydroxycarboxylic acid alone, however, inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid and the antimicrobial compound; (vi) No effect on metabolism in the presence of the 2-hydroxycarboxylic acid alone, however, an enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound. The composition according to claim 1.

7. The composition according to claim 1, wherein the dysregulation leads to sensitization of the bacteria to the antimicrobial compound.

8. The composition according to claim 1, wherein the 2-hydroxycarboxylic acid is D-lactic acid or a pharmaceutically acceptable salt thereof.

9. The composition according to claim 1, further comprising an antimicrobial compound.

10. A method for dysregulating the metabolism of planktonic bacteria, comprising the step of administering a composition containing 2-hydroxycarboxylic acid according to claim 1 or 2 to the planktonic bacteria.

11. A composition comprising a 2-hydroxycarboxylic acid according to claim 1 or 2 for use in treating or preventing a microbial infection in a subject, wherein the microbial infection is caused by a planktonic bacterium and the 2-hydroxycarboxylic acid dysregulates the metabolism of the planktonic bacterium.

12. Use of the 2-hydroxycarboxylic acid according to either claim 1 or 2 in the preparation of a composition for dysregulating the metabolism of planktonic bacteria.

13. Use of a composition comprising 2-hydroxycarboxylic acid according to claim 1 or 2 for dysregulating the metabolism of planktonic bacteria.

14. Use of a 2-hydroxycarboxylic acid according to either claim 1 or 2 in the manufacture of a composition for treating or preventing a microbial infection in a subject, wherein the microbial infection is caused by the planktonic bacteria and the composition dysregulates the metabolism of the planktonic bacteria.

15. Use of a composition comprising a 2-hydroxycarboxylic acid according to claim 1 or 2 for treating or preventing a microbial infection in a subject, wherein the microbial infection is caused by a planktonic bacterium and the 2-hydroxycarboxylic acid dysregulates the metabolism of the planktonic bacterium.

16. Use of a composition comprising 2-hydroxycarboxylic acid according to either claim 1 or 2 for dysregulating the metabolism of floating bacteria on a non-biological surface or in a non-biological substance.

17. A kit for dysregulating the metabolism of planktonic bacteria, wherein the kit is a) A composition comprising a 2-hydroxycarboxylic acid according to either claim 1 or 2; and b) Instructions for use A kit that includes this.

18. The aforementioned dysregulation, (i) Inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid alone, and inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (ii) Inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid alone, but enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (iii) Enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid alone, and enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (iv) Enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid alone, but inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound; (v) No effect on metabolism in the presence of the 2-hydroxycarboxylic acid alone, however, inhibition of metabolism in the presence of the 2-hydroxycarboxylic acid and the antimicrobial compound; (vi) No effect on metabolism in the presence of the 2-hydroxycarboxylic acid alone, however, an enhancement of metabolism in the presence of the 2-hydroxycarboxylic acid and an antimicrobial compound. The method according to claim 10, the use according to any one of claims 11 to 16, or the kit according to claim 17.

19. The method according to claim 10, the use according to any one of claims 11 to 16, or the kit according to claim 17, wherein the dysregulation leads to sensitization of the bacteria to the antimicrobial compound.

20. The method according to claim 10, the use according to any one of claims 11 to 16, or the kit according to claim 17, wherein the 2-hydroxycarboxylic acid is D-lactic acid or a pharmaceutically acceptable salt thereof.

21. The method according to claim 10, the use according to any one of claims 11 to 16, or the kit according to claim 17, wherein the composition further comprises an antimicrobial compound.

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